il 23  (Thermo Fisher)


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    Name:
    IL 23 Synthetic Peptide
    Description:
    IL 23 Synthetic Peptide for BLOCK Ctrl
    Catalog Number:
    PEP0357
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    Category:
    Proteins Enzymes Peptides
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    Structured Review

    Thermo Fisher il 23
    IL-1β and <t>IL-23</t> promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    IL 23 Synthetic Peptide for BLOCK Ctrl
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    il 23 - by Bioz Stars, 2021-06
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    Images

    1) Product Images from "Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression"

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression

    Journal: The FASEB Journal

    doi: 10.1096/fj.201802433R

    IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    Figure Legend Snippet: IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    2) Product Images from "Crassolide Suppresses Dendritic Cell Maturation and Attenuates Experimental Antiphospholipid Syndrome"

    Article Title: Crassolide Suppresses Dendritic Cell Maturation and Attenuates Experimental Antiphospholipid Syndrome

    Journal: Molecules

    doi: 10.3390/molecules26092492

    Effects of carssolide on the expression of surface markers of splenic DCs in APS mice. Spleen cells were purified from different mouse groups on day 56, and the relative mean fluorescence intensity (MFI) of ( a ) CD80 and ( b ) CD86 were measured by flow cytometry. Data are presented as mean ± SEM of 5 mice from one of three independent experiments. ( c ) IL-12 and ( d ) IL-23 mRNA expression levels in purified splenic CD11c+DCs determined by reverse transcription-quantitative PCR. Data were normalized to hypoxanthine guanine phosphoribosyl transferase 1 expression levels. * p
    Figure Legend Snippet: Effects of carssolide on the expression of surface markers of splenic DCs in APS mice. Spleen cells were purified from different mouse groups on day 56, and the relative mean fluorescence intensity (MFI) of ( a ) CD80 and ( b ) CD86 were measured by flow cytometry. Data are presented as mean ± SEM of 5 mice from one of three independent experiments. ( c ) IL-12 and ( d ) IL-23 mRNA expression levels in purified splenic CD11c+DCs determined by reverse transcription-quantitative PCR. Data were normalized to hypoxanthine guanine phosphoribosyl transferase 1 expression levels. * p

    Techniques Used: Expressing, Mouse Assay, Purification, Fluorescence, Flow Cytometry, Real-time Polymerase Chain Reaction

    3) Product Images from "Dendritic Cell IL-23 and IL-1 Production in Response to Schistosome Eggs Induces Th17 Cells in a Mouse Strain Prone to Severe Immunopathology 1"

    Article Title: Dendritic Cell IL-23 and IL-1 Production in Response to Schistosome Eggs Induces Th17 Cells in a Mouse Strain Prone to Severe Immunopathology 1

    Journal:

    doi:

    IL-17 production in DC-T cell cocultures stimulated with schistosome eggs or rIL-6, TGF- β , and IL-23. A–F , Naive CD4 T cells from CBA and BL/6 mice were cocultured with syngeneic BMDC in the presence or absence of anti-CD3/anti-CD28 coated
    Figure Legend Snippet: IL-17 production in DC-T cell cocultures stimulated with schistosome eggs or rIL-6, TGF- β , and IL-23. A–F , Naive CD4 T cells from CBA and BL/6 mice were cocultured with syngeneic BMDC in the presence or absence of anti-CD3/anti-CD28 coated

    Techniques Used: Crocin Bleaching Assay, Mouse Assay

    Neutralization of IL-23 and IL-1 inhibits egg-induced IL-17 production in CBA DC-T cell cocultures. A and B , DC-T cell cocultures were established in the presence or absence of eggs, anti-CD3/anti-CD28 coated beads, and the indicated blocking reagents
    Figure Legend Snippet: Neutralization of IL-23 and IL-1 inhibits egg-induced IL-17 production in CBA DC-T cell cocultures. A and B , DC-T cell cocultures were established in the presence or absence of eggs, anti-CD3/anti-CD28 coated beads, and the indicated blocking reagents

    Techniques Used: Neutralization, Crocin Bleaching Assay, Blocking Assay

    Exogenous IL-23 and IL-1 β enhance egg-induced IL-17 production in BL/6 DC-T cell cocultures. A–C , DC-T cell cocultures were established in the presence or absence of eggs, anti-CD3/anti-CD28 coated beads, and recombinant cytokines as described
    Figure Legend Snippet: Exogenous IL-23 and IL-1 β enhance egg-induced IL-17 production in BL/6 DC-T cell cocultures. A–C , DC-T cell cocultures were established in the presence or absence of eggs, anti-CD3/anti-CD28 coated beads, and recombinant cytokines as described

    Techniques Used: Recombinant

    4) Product Images from "Dissection of intercellular communication using the transcriptome-based framework ICELLNET"

    Article Title: Dissection of intercellular communication using the transcriptome-based framework ICELLNET

    Journal: Nature Communications

    doi: 10.1038/s41467-021-21244-x

    IL-10R blocking activates a cell-to-cell communication module in LPS-stimulated DCs. a Depicted are the four experimental conditions for which transcriptomics was generated ( n = 6 biologically independent samples). b Connectivity maps describing outward communication from DCs to putative target cells in the conditions: Medium (M), LPS (L), LPS + aTNFR and LPS + aIL-10R. Twelve primary cell types are considered as partner cells (DC: conventional dendritic cell, Macroph: macrophage, Mono: monocyte, Ostblast: osteoblast, Endoth: endothelial cell, Fblast: fibroblast, Kerat: keratinocyte, Epith: epithelial cell, B cell, T cell, Neutrop: neutrophil and NK: natural killer cell) and are colored depending on the cell compartment (green: stroma, orange: innate, blue: adaptive, pink: epithelium). The width of the edges corresponds to a global score combining the intensity of all the individual ligand/receptor interactions, normalized to the medium condition. A scale ranging from 1 to 10, corresponding to minimum and maximum communication scores, is shown in the legend. c Gene corresponding to ligands (black) and receptors (white) counted in each loop signature (from n = 6 biologically independent samples for each conditions) and plotted according to regulation directionality: upregulated (Up) or downregulated (Down). Genes with separability score ≥4 were included in each condition’s signature. d Balloon plot representing a selection of individual interaction scores. e Protein levels of IL-6 ( n = 10 biologically independent samples, except for Medium condition where n = 5), OSM ( n = 4 biologically independent samples), IL-23 ( n = 8 biologically independent samples) and IL-12p70 ( n = 16 biologically independent samples, except for Medium condition where n = 8) demonstrating increased secretion in LPS + aIL-10R DC supernatant. Data are presented as mean values ± SEM. Some schematic art pieces were used and modified from Servier Medical Art, licensed under a Creative Common Attribution 3.0 Generic License. http://smart.servier.com/ ( a ).
    Figure Legend Snippet: IL-10R blocking activates a cell-to-cell communication module in LPS-stimulated DCs. a Depicted are the four experimental conditions for which transcriptomics was generated ( n = 6 biologically independent samples). b Connectivity maps describing outward communication from DCs to putative target cells in the conditions: Medium (M), LPS (L), LPS + aTNFR and LPS + aIL-10R. Twelve primary cell types are considered as partner cells (DC: conventional dendritic cell, Macroph: macrophage, Mono: monocyte, Ostblast: osteoblast, Endoth: endothelial cell, Fblast: fibroblast, Kerat: keratinocyte, Epith: epithelial cell, B cell, T cell, Neutrop: neutrophil and NK: natural killer cell) and are colored depending on the cell compartment (green: stroma, orange: innate, blue: adaptive, pink: epithelium). The width of the edges corresponds to a global score combining the intensity of all the individual ligand/receptor interactions, normalized to the medium condition. A scale ranging from 1 to 10, corresponding to minimum and maximum communication scores, is shown in the legend. c Gene corresponding to ligands (black) and receptors (white) counted in each loop signature (from n = 6 biologically independent samples for each conditions) and plotted according to regulation directionality: upregulated (Up) or downregulated (Down). Genes with separability score ≥4 were included in each condition’s signature. d Balloon plot representing a selection of individual interaction scores. e Protein levels of IL-6 ( n = 10 biologically independent samples, except for Medium condition where n = 5), OSM ( n = 4 biologically independent samples), IL-23 ( n = 8 biologically independent samples) and IL-12p70 ( n = 16 biologically independent samples, except for Medium condition where n = 8) demonstrating increased secretion in LPS + aIL-10R DC supernatant. Data are presented as mean values ± SEM. Some schematic art pieces were used and modified from Servier Medical Art, licensed under a Creative Common Attribution 3.0 Generic License. http://smart.servier.com/ ( a ).

    Techniques Used: Blocking Assay, Generated, Selection, Modification

    5) Product Images from "Smad7 Controls Immunoregulatory PDL2/1-PD1 Signaling in Intestinal Inflammation and Autoimmunity"

    Article Title: Smad7 Controls Immunoregulatory PDL2/1-PD1 Signaling in Intestinal Inflammation and Autoimmunity

    Journal: Cell reports

    doi: 10.1016/j.celrep.2019.07.065

    Smad7 Deficiency in DCs Protects Mice against DSS-Induced Colitis (A) Percentage body weight changes (left) and linear regression analysis (right) of Smad7 fl/fl and DC-Smad7 −/− mice treated with 3% DSS in drinking water for 7 days (n = 8). (B) Disease activity index, a composite of body weight loss, blood in stool, and consistency of stool (higher score corresponds to colitis severity) (left) and linear regression analysis (right) of Smad7 fl/fl and DC-Smad7 −/− mice treated with 3% DSS in drinking water for 7 days (n = 9). (C) Representative histological sections with H E (left) and scores (n = 10) (right) of colitic mice based on degree of ulceration at day 7, scored blinded by a pathologist at HRHCF. Scale bars represent ~1 mm (top) and ~100 μm (bottom). (D) Colon lengths of colitic mice at day 7 (n = 13). (E) Representative NanoString (immunology panel) heatmap of inflammatory and regulatory genes in distal colon from colitic Smad7 fl/fl and DC-Smad7 −/− mice. Important upregulated and downregulated genes of interest in DC-Smad7 −/− are highlighted in red and green, respectively. (F) qRT-PCR validation of selected genes, including IL-1β, IL-6, TNF-α, and IL-18 expression in these samples (n = 3). (G) qRT-PCR of TGF-β, CD103, IL-12, IL-1β, IL-6, IL-23, and TNF-α expression in MLN CD11c + DCs from colitic mice (n = 5). (H) Representative FACS histograms (left) and MFIs (right) of PDL2, PDL1, and CD103 in MLN CD11c + DCs from colitic mice (n = 3). MFI data reflective of CD11c + population and expressed as FC from Smad7 fl/fl condition. (I) qRT-PCR of Foxp3, IFN-γ, and IL-17a in MLN CD4 + T cells (n = 5). (J) Representative FACS plots (left) and frequencies (right) of Foxp3 + , IFN-γ + , and IL-17a + populations in MLN (top) and lamina propria (LP) (bottom) CD4 + T cells (n = 4–10). (K) Percentage of body weight scores (left) and linear regression analyses (right) of Smad7 fl/fl and DC-Smad7 −/− mice treated during DSS administration, as indicated. Mice were treated with either immunoglobulin (Ig) control, (top) anti-CD25 (500 μg) (center), or anti-PD1 (250 μg) (bottom), given i.p. every other day 4 times, starting with DSS administration at day 0 (n = 3–4). Only control colitis body weight loss data for anti-CD25 are shown. Control data for anti-PD1 are identical (data not shown). Data representative of ≥3 independent experiments. qRT-PCR data expressed as FC from Smad7 fl/fl condition. Means ± SEMs. *p
    Figure Legend Snippet: Smad7 Deficiency in DCs Protects Mice against DSS-Induced Colitis (A) Percentage body weight changes (left) and linear regression analysis (right) of Smad7 fl/fl and DC-Smad7 −/− mice treated with 3% DSS in drinking water for 7 days (n = 8). (B) Disease activity index, a composite of body weight loss, blood in stool, and consistency of stool (higher score corresponds to colitis severity) (left) and linear regression analysis (right) of Smad7 fl/fl and DC-Smad7 −/− mice treated with 3% DSS in drinking water for 7 days (n = 9). (C) Representative histological sections with H E (left) and scores (n = 10) (right) of colitic mice based on degree of ulceration at day 7, scored blinded by a pathologist at HRHCF. Scale bars represent ~1 mm (top) and ~100 μm (bottom). (D) Colon lengths of colitic mice at day 7 (n = 13). (E) Representative NanoString (immunology panel) heatmap of inflammatory and regulatory genes in distal colon from colitic Smad7 fl/fl and DC-Smad7 −/− mice. Important upregulated and downregulated genes of interest in DC-Smad7 −/− are highlighted in red and green, respectively. (F) qRT-PCR validation of selected genes, including IL-1β, IL-6, TNF-α, and IL-18 expression in these samples (n = 3). (G) qRT-PCR of TGF-β, CD103, IL-12, IL-1β, IL-6, IL-23, and TNF-α expression in MLN CD11c + DCs from colitic mice (n = 5). (H) Representative FACS histograms (left) and MFIs (right) of PDL2, PDL1, and CD103 in MLN CD11c + DCs from colitic mice (n = 3). MFI data reflective of CD11c + population and expressed as FC from Smad7 fl/fl condition. (I) qRT-PCR of Foxp3, IFN-γ, and IL-17a in MLN CD4 + T cells (n = 5). (J) Representative FACS plots (left) and frequencies (right) of Foxp3 + , IFN-γ + , and IL-17a + populations in MLN (top) and lamina propria (LP) (bottom) CD4 + T cells (n = 4–10). (K) Percentage of body weight scores (left) and linear regression analyses (right) of Smad7 fl/fl and DC-Smad7 −/− mice treated during DSS administration, as indicated. Mice were treated with either immunoglobulin (Ig) control, (top) anti-CD25 (500 μg) (center), or anti-PD1 (250 μg) (bottom), given i.p. every other day 4 times, starting with DSS administration at day 0 (n = 3–4). Only control colitis body weight loss data for anti-CD25 are shown. Control data for anti-PD1 are identical (data not shown). Data representative of ≥3 independent experiments. qRT-PCR data expressed as FC from Smad7 fl/fl condition. Means ± SEMs. *p

    Techniques Used: Mouse Assay, Activity Assay, Quantitative RT-PCR, Expressing, FACS

    Smad7 Limits PD1 on CD4 + T Cells and PD1-Mediated Treg Differentiation and Promotes T Cell-Mediated Colitis (A) Representative FACS histograms (left) and MFIs (right) of phospho-Smad2/3 in naive CD4 + T cells from Smad7 fl/fl and T-Smad7 −/− mice stimulated with or without TGF-β (2.5 ng/mL), for 1 h (n = 4). (B) Representative FACS histograms (left) and MFIs (right) of PD1 in naive CD4 + T cells from these mice stimulated with low-dose plate-bound anti-CD3/CD28 (1 μg/mL) with or without TGF-β (2.5 ng/mL), for 18 h (n = 8). (C) Representative FACS plots (top), frequencies (bottom left), and cell counts per well (bottom right) of Foxp3 + populations in naive CD4 + T cells from these mice stimulated with or without TGF-β (0.5 ng/mL) and recombinant plate-bound PDL1-Fc or PDL2-Fc (30 μg/mL), for 4 days (n = 5). (D–H) Colitis was induced by adoptive transfer of 6 × 10 5 CD4 + CD45RB hi T cells i.p. into Rag-1 mice, followed by monitoring for 7 weeks. (D) Percentage body weight changes (left) and linear regression analysis (right) of Rag-1 −/− mice injected with CD4 + CD45RB hi T cells from Smad7 fl/fl or T-Smad7 −/− mice (n = 8). (E) Representative histological sections stained with H E of distal colons (left) and histological scores (n = 8) (right) from recipient Rag-1 −/− mice, based on the degree of epithelial damage, as scored blinded by a pathologist at HRHCF. Scale bars represent ~1 mm (top) and ~100 μm (bottom). (F) qRT-PCR of IL-12, IL-1β, IL-6, IL-23, and TNF-α in colitic tissue from recipient mice (n = 8). (G) Representative FACS plots (left) and frequencies (right) of Foxp3 + populations in MLN (top) and splenic (bottom) CD4 + T cells from recipient mice at 7 weeks (n = 5–8). (H) Representative FACS histograms (left) and frequencies (right) of PD1 in MLN (top) and splenic (bottom) CD4 + T cells from recipient mice at 1 week (n = 6). Data representative of ≥2 independent experiments. qRT-PCR and MFI (reflective of CD4 + population) data expressed as FC from Smad7 fl/fl media condition. Statistical comparisons to Smad7 fl/fl condition. Means ± SEMs. *p
    Figure Legend Snippet: Smad7 Limits PD1 on CD4 + T Cells and PD1-Mediated Treg Differentiation and Promotes T Cell-Mediated Colitis (A) Representative FACS histograms (left) and MFIs (right) of phospho-Smad2/3 in naive CD4 + T cells from Smad7 fl/fl and T-Smad7 −/− mice stimulated with or without TGF-β (2.5 ng/mL), for 1 h (n = 4). (B) Representative FACS histograms (left) and MFIs (right) of PD1 in naive CD4 + T cells from these mice stimulated with low-dose plate-bound anti-CD3/CD28 (1 μg/mL) with or without TGF-β (2.5 ng/mL), for 18 h (n = 8). (C) Representative FACS plots (top), frequencies (bottom left), and cell counts per well (bottom right) of Foxp3 + populations in naive CD4 + T cells from these mice stimulated with or without TGF-β (0.5 ng/mL) and recombinant plate-bound PDL1-Fc or PDL2-Fc (30 μg/mL), for 4 days (n = 5). (D–H) Colitis was induced by adoptive transfer of 6 × 10 5 CD4 + CD45RB hi T cells i.p. into Rag-1 mice, followed by monitoring for 7 weeks. (D) Percentage body weight changes (left) and linear regression analysis (right) of Rag-1 −/− mice injected with CD4 + CD45RB hi T cells from Smad7 fl/fl or T-Smad7 −/− mice (n = 8). (E) Representative histological sections stained with H E of distal colons (left) and histological scores (n = 8) (right) from recipient Rag-1 −/− mice, based on the degree of epithelial damage, as scored blinded by a pathologist at HRHCF. Scale bars represent ~1 mm (top) and ~100 μm (bottom). (F) qRT-PCR of IL-12, IL-1β, IL-6, IL-23, and TNF-α in colitic tissue from recipient mice (n = 8). (G) Representative FACS plots (left) and frequencies (right) of Foxp3 + populations in MLN (top) and splenic (bottom) CD4 + T cells from recipient mice at 7 weeks (n = 5–8). (H) Representative FACS histograms (left) and frequencies (right) of PD1 in MLN (top) and splenic (bottom) CD4 + T cells from recipient mice at 1 week (n = 6). Data representative of ≥2 independent experiments. qRT-PCR and MFI (reflective of CD4 + population) data expressed as FC from Smad7 fl/fl media condition. Statistical comparisons to Smad7 fl/fl condition. Means ± SEMs. *p

    Techniques Used: FACS, Mouse Assay, Recombinant, Adoptive Transfer Assay, Injection, Staining, Quantitative RT-PCR

    6) Product Images from "CD163+ cytokine-producing cDC2 stimulate intratumoral type 1 T cell responses in HPV16-induced oropharyngeal cancer"

    Article Title: CD163+ cytokine-producing cDC2 stimulate intratumoral type 1 T cell responses in HPV16-induced oropharyngeal cancer

    Journal: Journal for Immunotherapy of Cancer

    doi: 10.1136/jitc-2020-001053

    CD14 ‒ CD163 ‒ and CD14 ‒ CD163 + myeloid cells have allogeneic T cell stimulatory capacity and represent true DC. FACS-sorted CD14 + CD163 ‒ , CD14 + CD163 + , CD14 ‒ CD163 ‒ and CD14 ‒ CD163 + myeloid cells were tested for their T cell stimulatory and DC cytokine-producing potential. (A) Graphs depicting the percentage proliferation of CD4 + CD45RO + (left) and CD8 + CD45RO + (right) T cells within allogeneic PBMCs in response to CD14 + CD163 ‒ (open red), CD14 + CD163 + (closed red), CD14 ‒ CD163 ‒ (open blue) and CD14 ‒ CD163 + (closed blue) myeloid cells isolated from healthy donors (n=9, mean±SEM). (B) Graph depicting the IFNγ production in pg/mL of the total allogeneic PBMCs in response to the CD163 ‒ and CD163 + myeloid cells, as determined by ELISA (n=9, mean±SEM). (C) Graphs depicting the percentage of IFNγ + cells of proliferated CD4 + CD45RO + (left) and CD8 + CD45RO + (right) T cells in response to CD14 ‒ CD163 ‒ (open blue) and CD14 ‒ CD163 + (closed blue) myeloid cells isolated from healthy donors (n=7, mean±SEM). (D) Graphs depicting the IL-13, IL-9 and IL-22 production in pg/mL of the total allogeneic PBMCs in response to the CD163 ‒ and CD163 + myeloid cells (n=8, mean±SEM), as determined by multiplex T cell cytokine assay. The dotted line indicates the lower limit of detection of each of the cytokines. (E, F) Heatmaps presenting (E) IL-12p70 and IL-18 (n=15) and (F) Granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-1α, IL-1β, IL-10, IL-23, IL-6, IL-8, MIP-1α, MIP-3α, TGF-α, TNF-α and VEGF-A levels (n=12) in supernatants from toll-like receptor (TLR)-ligand stimulated CD14 + CD163 ‒ , CD14 + CD163 + , CD14 ‒ CD163 ‒ and CD14 ‒ CD163 + myeloid cells. Cytokines were determined by ELISA (E) and multiplex cytokine assays (F) and given as log10 values in pg/mL. *p
    Figure Legend Snippet: CD14 ‒ CD163 ‒ and CD14 ‒ CD163 + myeloid cells have allogeneic T cell stimulatory capacity and represent true DC. FACS-sorted CD14 + CD163 ‒ , CD14 + CD163 + , CD14 ‒ CD163 ‒ and CD14 ‒ CD163 + myeloid cells were tested for their T cell stimulatory and DC cytokine-producing potential. (A) Graphs depicting the percentage proliferation of CD4 + CD45RO + (left) and CD8 + CD45RO + (right) T cells within allogeneic PBMCs in response to CD14 + CD163 ‒ (open red), CD14 + CD163 + (closed red), CD14 ‒ CD163 ‒ (open blue) and CD14 ‒ CD163 + (closed blue) myeloid cells isolated from healthy donors (n=9, mean±SEM). (B) Graph depicting the IFNγ production in pg/mL of the total allogeneic PBMCs in response to the CD163 ‒ and CD163 + myeloid cells, as determined by ELISA (n=9, mean±SEM). (C) Graphs depicting the percentage of IFNγ + cells of proliferated CD4 + CD45RO + (left) and CD8 + CD45RO + (right) T cells in response to CD14 ‒ CD163 ‒ (open blue) and CD14 ‒ CD163 + (closed blue) myeloid cells isolated from healthy donors (n=7, mean±SEM). (D) Graphs depicting the IL-13, IL-9 and IL-22 production in pg/mL of the total allogeneic PBMCs in response to the CD163 ‒ and CD163 + myeloid cells (n=8, mean±SEM), as determined by multiplex T cell cytokine assay. The dotted line indicates the lower limit of detection of each of the cytokines. (E, F) Heatmaps presenting (E) IL-12p70 and IL-18 (n=15) and (F) Granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-1α, IL-1β, IL-10, IL-23, IL-6, IL-8, MIP-1α, MIP-3α, TGF-α, TNF-α and VEGF-A levels (n=12) in supernatants from toll-like receptor (TLR)-ligand stimulated CD14 + CD163 ‒ , CD14 + CD163 + , CD14 ‒ CD163 ‒ and CD14 ‒ CD163 + myeloid cells. Cytokines were determined by ELISA (E) and multiplex cytokine assays (F) and given as log10 values in pg/mL. *p

    Techniques Used: FACS, Isolation, Enzyme-linked Immunosorbent Assay, Multiplex Assay, Cytokine Assay

    7) Product Images from "Psoriatic skin inflammation is promoted by c‐Jun/AP‐1‐dependent CCL2 and IL‐23 expression in dendritic cells"

    Article Title: Psoriatic skin inflammation is promoted by c‐Jun/AP‐1‐dependent CCL2 and IL‐23 expression in dendritic cells

    Journal: EMBO Molecular Medicine

    doi: 10.15252/emmm.202012409

    c‐Jun directly regulates IL‐23 expression in DCs Heat map visualization of AP‐1 family member expression in DCs sorted from back skin of indicated mice after 4 h IMQ stimulation as quantified by qRT–PCR. Color code (red = up, blue = down‐regulation) shows fold change of Ap‐1 mRNA to untreated c‐Jun fl/fl DC control. Asterisk shows significant differences in IMQ activated to control c‐Jun fl/fl DCs ( n = 4–14; 2–4 independent experiments). Graph shows the percentage of c‐Jun positive skin DCs quantified by immunofluorescence performed as described in Fig EV2 (A) ( n = 5, 3 experiments). qRT–PCR detection of Il23p19 and Il17a in total RNA isolated from the back skin of indicated mice treated with IMQ for 32 or 48 h ( n = 10–21; 3–6 experiments). IL‐23 and IL‐17A protein was quantified by Luminex multiplex assay in cutaneous lysates of indicated mice after IMQ treatment (48 h) ( n = 5; 2 independent experiments). qRT–PCR detection of Il23p19 mRNA expression levels in BMDCs of indicated genotype stimulated with IMQ for 4 h and/or pretreated with JNKi (SP600125, 25 µM, 1 h) ( n = 4–14; 2–5 independent experiments). IL‐23 protein was quantified by ELISA (1 × 10 6 BMDCs/well) and by intracellular flow cytometry (IL‐23p19 + p40 + cells among live, single, CD45 + , CD11c + cells) in c‐Jun fl/fl and c‐Jun Δ/Δ Mx1 ‐Cre BMDCs stimulated with IMQ for 4 h (Left: n = 10–13; Right: n = 9–10; ≥ 3 independent experiments). qRT–PCR detection of Il23p19 mRNA expression levels in CD11c + MHCII + cells sorted from IMQ (8 h)‐treated back skin of indicated mice ( n = 5–12; 3 experiments). Genomic location and sequence of a putative AP‐1 binding site in the mouse promoter of IL‐23p19. Blockade of c‐Jun/AP‐1 TF binding by inhibition of phosphorylation (JNKi, red), or mutation of the binding sequence (mut 1 = T ‐ > C; green | mut 2 = TG ‐ > AT; blue) reduces IL‐23p19 promoter activity (see Fig 4J ). c‐Jun chromatin immunoprecipitation of c‐Jun fl/fl and c‐Jun Δ/Δ Mx1 ‐Cre BMDCs stimulated with IMQ for 2 h ( n = 4–6; 3 independent experiments). Luciferase activity was quantified in lysates of RAW 264.7 cells transfected with pGL3 basic vector harboring wild‐type (wt) or mutated (mut1 or 2) IL‐23p19 promoter. Transfected cells were pretreated with JNKi (SP600125, 25 µM, 1 h) and stimulated with IMQ (6 h). Results are shown as fold change to pGL‐3‐wt transfected, LAL (Limulus amebocyte lysate)‐treated RAW264.7 cells ( n = 4; 2 independent experiments). IL‐17A production by dermal γδ T cells was analyzed in IMQ‐treated skin (Day 2) of indicated mice by intracellular flow cytometry. Shown are representative plots (left) and a bar graph (right) of IL‐17A + cells pregated on live, single, CD45 + , γδ TCR int+ cells ( n = 7–10; 3 independent experiments). Epidermal thickness of back skin was quantified in H E stained sections of indicated mice treated with rIL‐23 (i.d., 1 µg) and/or IMQ for two consecutive days ( n = 4–7; 3 independent experiments). Data information: Data are shown as mean ± SEM. P ‐values were calculated by unpaired, two‐tailed t ‐test (A, C) and one‐way ANOVA with Tukey multiple comparison test (B, D‐G, I‐L). Statistical significance: ns > 0.05, * P
    Figure Legend Snippet: c‐Jun directly regulates IL‐23 expression in DCs Heat map visualization of AP‐1 family member expression in DCs sorted from back skin of indicated mice after 4 h IMQ stimulation as quantified by qRT–PCR. Color code (red = up, blue = down‐regulation) shows fold change of Ap‐1 mRNA to untreated c‐Jun fl/fl DC control. Asterisk shows significant differences in IMQ activated to control c‐Jun fl/fl DCs ( n = 4–14; 2–4 independent experiments). Graph shows the percentage of c‐Jun positive skin DCs quantified by immunofluorescence performed as described in Fig EV2 (A) ( n = 5, 3 experiments). qRT–PCR detection of Il23p19 and Il17a in total RNA isolated from the back skin of indicated mice treated with IMQ for 32 or 48 h ( n = 10–21; 3–6 experiments). IL‐23 and IL‐17A protein was quantified by Luminex multiplex assay in cutaneous lysates of indicated mice after IMQ treatment (48 h) ( n = 5; 2 independent experiments). qRT–PCR detection of Il23p19 mRNA expression levels in BMDCs of indicated genotype stimulated with IMQ for 4 h and/or pretreated with JNKi (SP600125, 25 µM, 1 h) ( n = 4–14; 2–5 independent experiments). IL‐23 protein was quantified by ELISA (1 × 10 6 BMDCs/well) and by intracellular flow cytometry (IL‐23p19 + p40 + cells among live, single, CD45 + , CD11c + cells) in c‐Jun fl/fl and c‐Jun Δ/Δ Mx1 ‐Cre BMDCs stimulated with IMQ for 4 h (Left: n = 10–13; Right: n = 9–10; ≥ 3 independent experiments). qRT–PCR detection of Il23p19 mRNA expression levels in CD11c + MHCII + cells sorted from IMQ (8 h)‐treated back skin of indicated mice ( n = 5–12; 3 experiments). Genomic location and sequence of a putative AP‐1 binding site in the mouse promoter of IL‐23p19. Blockade of c‐Jun/AP‐1 TF binding by inhibition of phosphorylation (JNKi, red), or mutation of the binding sequence (mut 1 = T ‐ > C; green | mut 2 = TG ‐ > AT; blue) reduces IL‐23p19 promoter activity (see Fig 4J ). c‐Jun chromatin immunoprecipitation of c‐Jun fl/fl and c‐Jun Δ/Δ Mx1 ‐Cre BMDCs stimulated with IMQ for 2 h ( n = 4–6; 3 independent experiments). Luciferase activity was quantified in lysates of RAW 264.7 cells transfected with pGL3 basic vector harboring wild‐type (wt) or mutated (mut1 or 2) IL‐23p19 promoter. Transfected cells were pretreated with JNKi (SP600125, 25 µM, 1 h) and stimulated with IMQ (6 h). Results are shown as fold change to pGL‐3‐wt transfected, LAL (Limulus amebocyte lysate)‐treated RAW264.7 cells ( n = 4; 2 independent experiments). IL‐17A production by dermal γδ T cells was analyzed in IMQ‐treated skin (Day 2) of indicated mice by intracellular flow cytometry. Shown are representative plots (left) and a bar graph (right) of IL‐17A + cells pregated on live, single, CD45 + , γδ TCR int+ cells ( n = 7–10; 3 independent experiments). Epidermal thickness of back skin was quantified in H E stained sections of indicated mice treated with rIL‐23 (i.d., 1 µg) and/or IMQ for two consecutive days ( n = 4–7; 3 independent experiments). Data information: Data are shown as mean ± SEM. P ‐values were calculated by unpaired, two‐tailed t ‐test (A, C) and one‐way ANOVA with Tukey multiple comparison test (B, D‐G, I‐L). Statistical significance: ns > 0.05, * P

    Techniques Used: Expressing, Mouse Assay, Quantitative RT-PCR, Immunofluorescence, Isolation, Luminex, Multiplex Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Sequencing, Binding Assay, Inhibition, Mutagenesis, Activity Assay, Chromatin Immunoprecipitation, Luciferase, Transfection, Plasmid Preparation, Staining, Two Tailed Test

    Characterization of c‐Jun co‐expression with CCL2 and IL‐23 in DCs of lesional psoriatic skin Immunofluorescence of c‐Jun (green), CD11c (red) and DAPI in psoriatic lesions from two patients and in non‐lesional skin from one patient. Insets give an enlarged view of the framed area. Arrows indicate CD11c + skin cells. Scale bar: 10 µm, Magnification: 40x. Linear regression analysis between c‐Jun and Ccl2 in healthy ( n = 38), non‐lesional ( n = 27) and lesional ( n = 28) skin. Expression values were obtained from the GEO Data Set [GSE121212]. A Pearson’s correlation test was performed and r 2 and P values are indicated in the plot. Expression values of Tlr7 , Tlr8 , Jnk1 , Jnk2 , Jnk3, c‐Jun, Ccl2 and Il23p19 in healthy ( n = 38), non‐lesional ( n = 27) and lesional ( n = 28) skin. Expression values were obtained from the GEO Data Set [GSE121212]. Box and whiskers plot: Central band shows median, box extends from the 25 th to 75 th percentiles and whiskers go down to the smallest (min) and up to the largest (max) value. Representative immunofluorescence of CCL2 or IL‐23p19 (green) and DAPI in healthy and lesional skin. Arrows indicate IL‐23p19 + (upper panels) or CCL2 + dermal cells (lower panels). Magnification: 40x. Scale bar: 50 µm ( n = 2 patient samples). Representative immunofluorescence of CCL2 (E) or IL‐23p19 (F) (white), c‐Jun (green) and the DC markers CD1c or CD14 (red) was performed on lesional psoriatic skin. DAPI was used to counterstain. Arrows indicate triple‐positive cells and an asterisk highlights a representative one that is shown enlarged in a white‐framed inlet. Magnification: 40×. Scale bar: 50 µm ( n = 2 patient samples). Data information: Data are shown as mean ± SEM. P ‐values were calculated by Pearson´s correlation test (B) and one‐way ANOVA with Tukey multiple comparison test (C). Statistical significance: ns > 0.05, ** P
    Figure Legend Snippet: Characterization of c‐Jun co‐expression with CCL2 and IL‐23 in DCs of lesional psoriatic skin Immunofluorescence of c‐Jun (green), CD11c (red) and DAPI in psoriatic lesions from two patients and in non‐lesional skin from one patient. Insets give an enlarged view of the framed area. Arrows indicate CD11c + skin cells. Scale bar: 10 µm, Magnification: 40x. Linear regression analysis between c‐Jun and Ccl2 in healthy ( n = 38), non‐lesional ( n = 27) and lesional ( n = 28) skin. Expression values were obtained from the GEO Data Set [GSE121212]. A Pearson’s correlation test was performed and r 2 and P values are indicated in the plot. Expression values of Tlr7 , Tlr8 , Jnk1 , Jnk2 , Jnk3, c‐Jun, Ccl2 and Il23p19 in healthy ( n = 38), non‐lesional ( n = 27) and lesional ( n = 28) skin. Expression values were obtained from the GEO Data Set [GSE121212]. Box and whiskers plot: Central band shows median, box extends from the 25 th to 75 th percentiles and whiskers go down to the smallest (min) and up to the largest (max) value. Representative immunofluorescence of CCL2 or IL‐23p19 (green) and DAPI in healthy and lesional skin. Arrows indicate IL‐23p19 + (upper panels) or CCL2 + dermal cells (lower panels). Magnification: 40x. Scale bar: 50 µm ( n = 2 patient samples). Representative immunofluorescence of CCL2 (E) or IL‐23p19 (F) (white), c‐Jun (green) and the DC markers CD1c or CD14 (red) was performed on lesional psoriatic skin. DAPI was used to counterstain. Arrows indicate triple‐positive cells and an asterisk highlights a representative one that is shown enlarged in a white‐framed inlet. Magnification: 40×. Scale bar: 50 µm ( n = 2 patient samples). Data information: Data are shown as mean ± SEM. P ‐values were calculated by Pearson´s correlation test (B) and one‐way ANOVA with Tukey multiple comparison test (C). Statistical significance: ns > 0.05, ** P

    Techniques Used: Expressing, Immunofluorescence

    Role of JNK‐AP‐1 signaling in the maturation of human mo‐DCs stimulated with R848 or LL‐37‐RNA‐40 Human mo‐DCs were pretreated with DMSO (1:1,000) or JNK Inhibitor (SP600125, 25 µM) (A, B) or AP‐1 Inhibitor (T‐5224, 20 µM) (C, D) for 1 h and stimulated with R848 for 24 h. Gated CD1a + cells were analyzed for surface expression of CD80 and CD86 by flow cytometry. Shown are representative histograms (A, C) and mean fluorescence intensity (MFI) (B, D) ( n = 4 (B); representative result from 2 independent experiments is shown, n = 7 (D); 2 independent experiments). Human mo‐DCs were stimulated with LL‐37‐RNA‐40 complex or LL‐37 or RNA‐40 for 24 h. Representative flow cytometry plots and histograms are shown in (E) and mean fluorescence intensity (MFI) of CD80 and C86 is shown in (F). Expression of CCL2 and IL‐23 was analyzed by ELISA (G) ( n = 7, 2 independent experiments). Data information: Data are shown as mean ± SEM. P ‐values were calculated by paired, two‐tailed t ‐test (B, D) or one‐way ANOVA with Tukey multiple comparison test (F, G). Statistical significance: ns > 0.05, * P
    Figure Legend Snippet: Role of JNK‐AP‐1 signaling in the maturation of human mo‐DCs stimulated with R848 or LL‐37‐RNA‐40 Human mo‐DCs were pretreated with DMSO (1:1,000) or JNK Inhibitor (SP600125, 25 µM) (A, B) or AP‐1 Inhibitor (T‐5224, 20 µM) (C, D) for 1 h and stimulated with R848 for 24 h. Gated CD1a + cells were analyzed for surface expression of CD80 and CD86 by flow cytometry. Shown are representative histograms (A, C) and mean fluorescence intensity (MFI) (B, D) ( n = 4 (B); representative result from 2 independent experiments is shown, n = 7 (D); 2 independent experiments). Human mo‐DCs were stimulated with LL‐37‐RNA‐40 complex or LL‐37 or RNA‐40 for 24 h. Representative flow cytometry plots and histograms are shown in (E) and mean fluorescence intensity (MFI) of CD80 and C86 is shown in (F). Expression of CCL2 and IL‐23 was analyzed by ELISA (G) ( n = 7, 2 independent experiments). Data information: Data are shown as mean ± SEM. P ‐values were calculated by paired, two‐tailed t ‐test (B, D) or one‐way ANOVA with Tukey multiple comparison test (F, G). Statistical significance: ns > 0.05, * P

    Techniques Used: Expressing, Flow Cytometry, Fluorescence, Enzyme-linked Immunosorbent Assay, Two Tailed Test

    Validation of the therapeutic potential of JNK inhibition in psoriasis‐like mouse models Back skin of c‐Jun fl/fl , c‐Jun Δ/Δ CD11c ‐Cre, Tlr7 −/− and Tlr7 −/− c‐Jun Δ/Δ CD11c ‐Cre mice was treated with IMQ and JNKi or vehicle for 5 days. Shown are representative H E stained sections (Bright‐field images, Magnification: 20×, Scale bar: 100 µm) (A), immunofluorescence staining’s of Ki‐67 (green) and DAPI (Scale bar: 100 µm, Magnification: 20×) (B) and of K5 (red), K10 (green) and DAPI (Scale bar: 100 µm, Magnification: 20×) (C) in murine back skin. For quantification 2–3 randomly chosen fields per section were counted. Flow cytometric analysis of cell suspension from back skin of indicated mice and treatment. t‐SNE‐ plots (D) show the cutaneous immune cell phenotype. Populations defined by t‐SNE algorithm were confirmed by conventional gating as described in Appendix Fig S7A and representative plots are shown in (E) for Monocytes and Neutrophils and in (F) for γδ T cells and dendritic epidermal T cells (DETC). Numbers adjacent to marked areas indicate percentage of cell population among CD45 + cells (D) and live, single cells (E, F). Representative plots shown are pregated on live, single, CD45 + (D) and CD11b + (E) or CD11b − cells (F). Experimental design for blocking IL‐23 signaling in the IMQ‐induced skin inflammation model. Back skin of wild‐type mice was treated daily with IMQ and anti‐IL23R antibody (15 mg/kg, i.p.) or isotype control for 5 consecutive days. To compare JNK Inhibitor (15 mg/kg, i.p.) was given ( n = 3) Trans‐epidermal water loss (TEWL) was analyzed in the back skin at the end of treatment. Flow cytometry of total back skin after 5 days of IMQ treatment. Analyzed were dermal γδ T cells (γδ TCR int+ ), Monocytes (CD11b + Ly6C hi ), and Neutrophils (CD11b + Ly6G + ). Graphs show immune cells as % of live, single cells. Data information: Data are shown as mean ± SEM. P ‐values were calculated by Tukey multiple comparison test (H, I). Statistical significance: ns > 0.05, * P
    Figure Legend Snippet: Validation of the therapeutic potential of JNK inhibition in psoriasis‐like mouse models Back skin of c‐Jun fl/fl , c‐Jun Δ/Δ CD11c ‐Cre, Tlr7 −/− and Tlr7 −/− c‐Jun Δ/Δ CD11c ‐Cre mice was treated with IMQ and JNKi or vehicle for 5 days. Shown are representative H E stained sections (Bright‐field images, Magnification: 20×, Scale bar: 100 µm) (A), immunofluorescence staining’s of Ki‐67 (green) and DAPI (Scale bar: 100 µm, Magnification: 20×) (B) and of K5 (red), K10 (green) and DAPI (Scale bar: 100 µm, Magnification: 20×) (C) in murine back skin. For quantification 2–3 randomly chosen fields per section were counted. Flow cytometric analysis of cell suspension from back skin of indicated mice and treatment. t‐SNE‐ plots (D) show the cutaneous immune cell phenotype. Populations defined by t‐SNE algorithm were confirmed by conventional gating as described in Appendix Fig S7A and representative plots are shown in (E) for Monocytes and Neutrophils and in (F) for γδ T cells and dendritic epidermal T cells (DETC). Numbers adjacent to marked areas indicate percentage of cell population among CD45 + cells (D) and live, single cells (E, F). Representative plots shown are pregated on live, single, CD45 + (D) and CD11b + (E) or CD11b − cells (F). Experimental design for blocking IL‐23 signaling in the IMQ‐induced skin inflammation model. Back skin of wild‐type mice was treated daily with IMQ and anti‐IL23R antibody (15 mg/kg, i.p.) or isotype control for 5 consecutive days. To compare JNK Inhibitor (15 mg/kg, i.p.) was given ( n = 3) Trans‐epidermal water loss (TEWL) was analyzed in the back skin at the end of treatment. Flow cytometry of total back skin after 5 days of IMQ treatment. Analyzed were dermal γδ T cells (γδ TCR int+ ), Monocytes (CD11b + Ly6C hi ), and Neutrophils (CD11b + Ly6G + ). Graphs show immune cells as % of live, single cells. Data information: Data are shown as mean ± SEM. P ‐values were calculated by Tukey multiple comparison test (H, I). Statistical significance: ns > 0.05, * P

    Techniques Used: Inhibition, Mouse Assay, Staining, Immunofluorescence, Blocking Assay, Flow Cytometry

    JNK/AP‐1 Inhibitors repress CCL2 and IL‐23 expression in human mo‐DCs Western blot analyses of Phospho‐ or total JNK and c‐Jun in human mo‐DCs pretreated with DMSO (1: 1,000) or SP600125 (JNKi, 25 µM) for 1 h and stimulated with R848 (10 µg/mL) for 15 min. Human mo‐DCs pretreated with JNKi (SP600125, 25 µM) (B) or AP‐1 Inhibitor (T‐5224, 20 µM) (C) for 1 h were stimulated with R848 (10 µg/ml). CCL2 and IL‐23 were analyzed by ELISA ( n = 5–11; 2–3 independent experiments). Representative flow cytometry plots (left) and histograms (right) of human mo‐DCs stimulated with LL‐37‐RNA‐40 for 24 h are shown. Inhibitors (JNKi; 25 µM or AP‐1 Inhibitor; 20 µM) were given 1 h before stimulation. Plots shown are pregated on live, single, CD1a + cells. Human mo‐DCs described in (D) were analyzed for mean fluorescence intensity of CD80 or CD86 by flow cytometry (E) and CCL2 or IL‐23 protein by ELISA (F) ( n = 7; 2 independent experiments). Data information: Data are shown as mean ± SEM. P ‐values were calculated by paired, two‐tailed t ‐test (B, C, E and F). Statistical significance: ns > 0.05, * P
    Figure Legend Snippet: JNK/AP‐1 Inhibitors repress CCL2 and IL‐23 expression in human mo‐DCs Western blot analyses of Phospho‐ or total JNK and c‐Jun in human mo‐DCs pretreated with DMSO (1: 1,000) or SP600125 (JNKi, 25 µM) for 1 h and stimulated with R848 (10 µg/mL) for 15 min. Human mo‐DCs pretreated with JNKi (SP600125, 25 µM) (B) or AP‐1 Inhibitor (T‐5224, 20 µM) (C) for 1 h were stimulated with R848 (10 µg/ml). CCL2 and IL‐23 were analyzed by ELISA ( n = 5–11; 2–3 independent experiments). Representative flow cytometry plots (left) and histograms (right) of human mo‐DCs stimulated with LL‐37‐RNA‐40 for 24 h are shown. Inhibitors (JNKi; 25 µM or AP‐1 Inhibitor; 20 µM) were given 1 h before stimulation. Plots shown are pregated on live, single, CD1a + cells. Human mo‐DCs described in (D) were analyzed for mean fluorescence intensity of CD80 or CD86 by flow cytometry (E) and CCL2 or IL‐23 protein by ELISA (F) ( n = 7; 2 independent experiments). Data information: Data are shown as mean ± SEM. P ‐values were calculated by paired, two‐tailed t ‐test (B, C, E and F). Statistical significance: ns > 0.05, * P

    Techniques Used: Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Fluorescence, Two Tailed Test

    c‐Jun, CCL2, and IL‐23 are co‐expressed in type‐2/inflammatory DCs in psoriatic lesions Gene expression values of Cd207 , Cd1a , Cd1c , Cd14 , Xcr1 in healthy ( n = 38), non‐lesional ( n = 27), and lesional ( n = 28) skin. Expression values were obtained from the GEO Data Set [GSE121212]. Box and whiskers plot: Central band shows median, box extends from the 25 th to 75 th percentiles, and whiskers go down to the smallest (min) and up to the largest (max) value. Immunofluorescence of c‐Jun (green), CD1a or CD1c or CD14 (red) and DAPI in psoriatic lesions. Shown is a representative image (Magnification: 25×, Scale Bar: 100µm) with an Inset (Magnification: 63×, Scale Bar: 20 µm, Deconvoluted). Arrows indicate c‐Jun + DCs. Asterisk highlights a DC with prominent c‐Jun expression as shown enlarged in 2 white‐framed windows (DC Marker (red) + c‐Jun (green, left) or DAPI (blue, right)). Quantification of c‐Jun + DCs (CD1a, CD1c, or CD14) in psoriatic lesions as shown in (B). Four randomly chosen fields on one section were analyzed for each sample ( n = 2 patient samples/condition). Box and whiskers plot: Central band shows median, box extends from the 25 th to 75 th percentiles, and whiskers go down to the smallest (min) and up to the largest (max) value. Representative immunofluorescence of c‐Jun (green), CD1a (red), CCL2 (D), or IL‐23p19 (E) (white) and DAPI in psoriatic lesions. Arrows indicate triple‐positive cells and an asterisk highlights a representative one that is shown enlarged in a white‐framed inlet. Magnification: 40x. Scale bar: 50 µm ( n = 2 patient samples). Data information: Data are shown as mean ± SEM. P ‐values were calculated by one‐way ANOVA with Tukey multiple comparison test (A, C). Statistical significance: ns > 0.05, * P
    Figure Legend Snippet: c‐Jun, CCL2, and IL‐23 are co‐expressed in type‐2/inflammatory DCs in psoriatic lesions Gene expression values of Cd207 , Cd1a , Cd1c , Cd14 , Xcr1 in healthy ( n = 38), non‐lesional ( n = 27), and lesional ( n = 28) skin. Expression values were obtained from the GEO Data Set [GSE121212]. Box and whiskers plot: Central band shows median, box extends from the 25 th to 75 th percentiles, and whiskers go down to the smallest (min) and up to the largest (max) value. Immunofluorescence of c‐Jun (green), CD1a or CD1c or CD14 (red) and DAPI in psoriatic lesions. Shown is a representative image (Magnification: 25×, Scale Bar: 100µm) with an Inset (Magnification: 63×, Scale Bar: 20 µm, Deconvoluted). Arrows indicate c‐Jun + DCs. Asterisk highlights a DC with prominent c‐Jun expression as shown enlarged in 2 white‐framed windows (DC Marker (red) + c‐Jun (green, left) or DAPI (blue, right)). Quantification of c‐Jun + DCs (CD1a, CD1c, or CD14) in psoriatic lesions as shown in (B). Four randomly chosen fields on one section were analyzed for each sample ( n = 2 patient samples/condition). Box and whiskers plot: Central band shows median, box extends from the 25 th to 75 th percentiles, and whiskers go down to the smallest (min) and up to the largest (max) value. Representative immunofluorescence of c‐Jun (green), CD1a (red), CCL2 (D), or IL‐23p19 (E) (white) and DAPI in psoriatic lesions. Arrows indicate triple‐positive cells and an asterisk highlights a representative one that is shown enlarged in a white‐framed inlet. Magnification: 40x. Scale bar: 50 µm ( n = 2 patient samples). Data information: Data are shown as mean ± SEM. P ‐values were calculated by one‐way ANOVA with Tukey multiple comparison test (A, C). Statistical significance: ns > 0.05, * P

    Techniques Used: Expressing, Immunofluorescence, Marker

    c‐Jun is essential in conventional type‐2 DCs to control CCL2 and IL‐23 expression DC subsets (cDC1, cDC2), macrophages (MP), and Langerhans cells (LC) were sorted from IMQ‐treated back skin (5 h). c‐Jun , Ccl2, and Il23p19 mRNA was analyzed by qRT–PCR. Sort strategy is shown in (B) (10 mice were pooled per condition, 1 experiment). Sort strategy for the separation of MPs (CD45 + Lin − CD64 + CD172a + CD11b + ), and the DC (CD45 + Lin − CD64 − CD11c + MHCII + ) subsets cDC1 (XCR1 + CD11b − DC), cDC2 (XCR1 − CD11b + CD24 −/int CD326 − ) and LC (XCR1 − CD11b + CD24 + CD326 + ) in IMQ‐treated (5 h) murine back skin. qRT–PCR detection of c‐Jun , Ccl2, and Il23p19 in MPs and cDC2 sorted as described in (B) from indicated mice (MPs: n = 7–9; 3 independent experiments, cDC2: n = 5–10; 2–3 independent experiments and back skin from 2 mice was pooled). Sort strategy for CD11c expressing MPs (CD45 + Lin − CD11c int/low MHCII + CD64 + CD11b + CD172a + ) is shown. Murine skin was treated for 5 h with IMQ. Expression of indicated targets was analyzed by qRT–PCR in cells sorted as described in (D) ( n = 3; Back skin from 2 mice was pooled). Data information: Data are shown as mean ± SEM. P ‐values were calculated by unpaired, two‐tailed t ‐test (C, E). Statistical significance: ns > 0.05, * P
    Figure Legend Snippet: c‐Jun is essential in conventional type‐2 DCs to control CCL2 and IL‐23 expression DC subsets (cDC1, cDC2), macrophages (MP), and Langerhans cells (LC) were sorted from IMQ‐treated back skin (5 h). c‐Jun , Ccl2, and Il23p19 mRNA was analyzed by qRT–PCR. Sort strategy is shown in (B) (10 mice were pooled per condition, 1 experiment). Sort strategy for the separation of MPs (CD45 + Lin − CD64 + CD172a + CD11b + ), and the DC (CD45 + Lin − CD64 − CD11c + MHCII + ) subsets cDC1 (XCR1 + CD11b − DC), cDC2 (XCR1 − CD11b + CD24 −/int CD326 − ) and LC (XCR1 − CD11b + CD24 + CD326 + ) in IMQ‐treated (5 h) murine back skin. qRT–PCR detection of c‐Jun , Ccl2, and Il23p19 in MPs and cDC2 sorted as described in (B) from indicated mice (MPs: n = 7–9; 3 independent experiments, cDC2: n = 5–10; 2–3 independent experiments and back skin from 2 mice was pooled). Sort strategy for CD11c expressing MPs (CD45 + Lin − CD11c int/low MHCII + CD64 + CD11b + CD172a + ) is shown. Murine skin was treated for 5 h with IMQ. Expression of indicated targets was analyzed by qRT–PCR in cells sorted as described in (D) ( n = 3; Back skin from 2 mice was pooled). Data information: Data are shown as mean ± SEM. P ‐values were calculated by unpaired, two‐tailed t ‐test (C, E). Statistical significance: ns > 0.05, * P

    Techniques Used: Expressing, Quantitative RT-PCR, Mouse Assay, Two Tailed Test

    8) Product Images from "GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis"

    Article Title: GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.01520

    GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p
    Figure Legend Snippet: GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p

    Techniques Used: Recombinant, Enzyme-linked Immunosorbent Assay

    9) Product Images from "MiR-223 plays a protecting role in neutrophilic asthmatic mice through the inhibition of NLRP3 inflammasome"

    Article Title: MiR-223 plays a protecting role in neutrophilic asthmatic mice through the inhibition of NLRP3 inflammasome

    Journal: Respiratory Research

    doi: 10.1186/s12931-020-01374-4

    Blockade of NLRP3 inflammasome or IL-1β abrogates airway inflammation in miR-223 −/− mice. OVA/CFA was administrated to WT mice and miR-223 −/− mice on day 0. miR-223 −/− mice received the inhibitor of NLRP3, MCC950 (200 mg/kg, i.p.) and IL-1β receptor antagonist, anakinra (50 mg/kg, i.p.) or PBS after each challenge, respectively. Mice were euthanized 24 h after the final treatment. a Representative micrographs of lung H E staining a - d and PAS staining e - h from different groups after treatment (200× magnification). b - c Semi-quantification of lung inflammatory score and Apas + /Pbm was performed. n = 6–8 mice/group; statistical significance was determined by ANOVA. d Number of total inflammatory cells, neutrophils, eosinophils, and lymphocytes was calculated in BALF after treatment. n = 6–8 mice/group; statistical significance was determined by ANOVA. e AHR was determined by lung resistance after treatment. n = 6–8 mice/group. Statistical significance was determined by ANOVA. f - n The levels of Th2-associated cytokines (IL-4, IL-5, IL-13), Th17-associated cytokines (IL-17A, IL-22, IL-23), Th1-associated cytokine (IFN-γ), IL-1β, and IL-18 in BALF were measured by ELISA after treatment. n = 6–8 mice/group; statistical significance was determined by ANOVA. All data were expressed as mean ± SD. * P
    Figure Legend Snippet: Blockade of NLRP3 inflammasome or IL-1β abrogates airway inflammation in miR-223 −/− mice. OVA/CFA was administrated to WT mice and miR-223 −/− mice on day 0. miR-223 −/− mice received the inhibitor of NLRP3, MCC950 (200 mg/kg, i.p.) and IL-1β receptor antagonist, anakinra (50 mg/kg, i.p.) or PBS after each challenge, respectively. Mice were euthanized 24 h after the final treatment. a Representative micrographs of lung H E staining a - d and PAS staining e - h from different groups after treatment (200× magnification). b - c Semi-quantification of lung inflammatory score and Apas + /Pbm was performed. n = 6–8 mice/group; statistical significance was determined by ANOVA. d Number of total inflammatory cells, neutrophils, eosinophils, and lymphocytes was calculated in BALF after treatment. n = 6–8 mice/group; statistical significance was determined by ANOVA. e AHR was determined by lung resistance after treatment. n = 6–8 mice/group. Statistical significance was determined by ANOVA. f - n The levels of Th2-associated cytokines (IL-4, IL-5, IL-13), Th17-associated cytokines (IL-17A, IL-22, IL-23), Th1-associated cytokine (IFN-γ), IL-1β, and IL-18 in BALF were measured by ELISA after treatment. n = 6–8 mice/group; statistical significance was determined by ANOVA. All data were expressed as mean ± SD. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    MiR-223 played an important role in regulating neutrophilic airway inflammation in the OVA/CFA-induced asthma model. a Mice were sensitized against OVA in the presence of complete Freund’s adjuvant (CFA), followed by exposure to 1% OVA aerosols on days 21 and 22. Administration of miR-223 agomirs or negative control agomirs to OVA-sensitized mice for 3 consecutive days was shown. Analysis was performed at day 23. b MiR-223 expression levels were detected in the lung tissues from mice after OVA sensitization (1 day), after first challenged (22 day) and in asthma model (23 day) by qPCR, respectively. n = 6–8 mice/group; statistical significance was determined by ANOVA. c Representative micrographs of lung H E staining a - d and PAS staining e-h from different groups (200× magnification). d, e Semi-quantification of lung inflammatory score and Apas + /Pbm was performed. n = 6–8 mice/group; statistical significance was determined by ANOVA. f Number of total inflammatory cells, neutrophils, eosinophils, and lymphocytes was calculated in BALF. n = 6–8 mice/group; statistical significance was determined by ANOVA. g AHR was determined by lung resistance. n = 6–8 mice/group. Statistical significance was determined by ANOVA. h - p The levels of Th2-associated cytokines (IL-4, IL-5, IL-13), Th17-associated cytokines (IL-17A, IL-22, IL-23), Th1-associated cytokine (IFN-γ), IL-1β, and IL-18 and in BALF were measured by ELISA. n = 6–8 mice/group; statistical significance was determined by ANOVA. All data were expressed as mean ± SD. * P
    Figure Legend Snippet: MiR-223 played an important role in regulating neutrophilic airway inflammation in the OVA/CFA-induced asthma model. a Mice were sensitized against OVA in the presence of complete Freund’s adjuvant (CFA), followed by exposure to 1% OVA aerosols on days 21 and 22. Administration of miR-223 agomirs or negative control agomirs to OVA-sensitized mice for 3 consecutive days was shown. Analysis was performed at day 23. b MiR-223 expression levels were detected in the lung tissues from mice after OVA sensitization (1 day), after first challenged (22 day) and in asthma model (23 day) by qPCR, respectively. n = 6–8 mice/group; statistical significance was determined by ANOVA. c Representative micrographs of lung H E staining a - d and PAS staining e-h from different groups (200× magnification). d, e Semi-quantification of lung inflammatory score and Apas + /Pbm was performed. n = 6–8 mice/group; statistical significance was determined by ANOVA. f Number of total inflammatory cells, neutrophils, eosinophils, and lymphocytes was calculated in BALF. n = 6–8 mice/group; statistical significance was determined by ANOVA. g AHR was determined by lung resistance. n = 6–8 mice/group. Statistical significance was determined by ANOVA. h - p The levels of Th2-associated cytokines (IL-4, IL-5, IL-13), Th17-associated cytokines (IL-17A, IL-22, IL-23), Th1-associated cytokine (IFN-γ), IL-1β, and IL-18 and in BALF were measured by ELISA. n = 6–8 mice/group; statistical significance was determined by ANOVA. All data were expressed as mean ± SD. * P

    Techniques Used: Mouse Assay, Negative Control, Expressing, Real-time Polymerase Chain Reaction, Staining, Enzyme-linked Immunosorbent Assay

    MiR-223 agomirs attenuated airway inflammation in neutrophilic asthma. OVA/CFA was administrated to WT mice on day 0. Mice were treated with 5 nmol miR-223 agomirs or negative control agomirs on days 21–23 before each challenge, respectively. a The expression of miR-223 was detected in lung tissues by qPCR. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. b Representative micrographs of lung H E staining a - b and PAS staining c - d from treated mice (200× magnification). c - d Semi-quantification of lung inflammatory score and Apas + /Pbm was performed. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. e Number of total inflammatory cells, neutrophils, eosinophils, and lymphocytes was calculated in BALF after treatment. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. f AHR was determined by lung resistance after treatment. n = 6–8 mice/group. Statistical significance was determined by unpaired Student’s t test. g - o The levels of Th2-associated cytokines (IL-4, IL-5, IL-13), Th17-associated cytokines (IL-17A, IL-22, IL-23), Th1-associated cytokine (IFN-γ), IL-1β, and IL-18 in BALF were measured by ELISA after treatment. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. p Proteins expression of NLRP3, ASC, caspase-1, pro-caspase-1 and GAPDH were measured by western blot after treatment. Representative immunoblot from three independent mice. q Proteins expression of IL-1β, pro-IL-1β, IL-18, pro-IL-18, and GAPDH were measured using western blotting after treatment. Representative immunoblot from three independent mice. n = 6–8 mice/group; statistical significance was determined using ANOVA. r Genes expression of NLRP3, ASC, caspase-1, IL-1β and IL-18 were determined by qPCR after treatment. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. All data were expressed as mean ± SD. * P
    Figure Legend Snippet: MiR-223 agomirs attenuated airway inflammation in neutrophilic asthma. OVA/CFA was administrated to WT mice on day 0. Mice were treated with 5 nmol miR-223 agomirs or negative control agomirs on days 21–23 before each challenge, respectively. a The expression of miR-223 was detected in lung tissues by qPCR. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. b Representative micrographs of lung H E staining a - b and PAS staining c - d from treated mice (200× magnification). c - d Semi-quantification of lung inflammatory score and Apas + /Pbm was performed. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. e Number of total inflammatory cells, neutrophils, eosinophils, and lymphocytes was calculated in BALF after treatment. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. f AHR was determined by lung resistance after treatment. n = 6–8 mice/group. Statistical significance was determined by unpaired Student’s t test. g - o The levels of Th2-associated cytokines (IL-4, IL-5, IL-13), Th17-associated cytokines (IL-17A, IL-22, IL-23), Th1-associated cytokine (IFN-γ), IL-1β, and IL-18 in BALF were measured by ELISA after treatment. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. p Proteins expression of NLRP3, ASC, caspase-1, pro-caspase-1 and GAPDH were measured by western blot after treatment. Representative immunoblot from three independent mice. q Proteins expression of IL-1β, pro-IL-1β, IL-18, pro-IL-18, and GAPDH were measured using western blotting after treatment. Representative immunoblot from three independent mice. n = 6–8 mice/group; statistical significance was determined using ANOVA. r Genes expression of NLRP3, ASC, caspase-1, IL-1β and IL-18 were determined by qPCR after treatment. n = 6–8 mice/group; statistical significance was determined by unpaired Student’s t test. All data were expressed as mean ± SD. * P

    Techniques Used: Mouse Assay, Negative Control, Expressing, Real-time Polymerase Chain Reaction, Staining, Enzyme-linked Immunosorbent Assay, Western Blot

    10) Product Images from "Characterization of the IL-17 and CD4+ Th17 Cells in the Clinical Course of Dengue Virus Infections"

    Article Title: Characterization of the IL-17 and CD4+ Th17 Cells in the Clinical Course of Dengue Virus Infections

    Journal: Viruses

    doi: 10.3390/v12121435

    Serum levels of IL-17A and IL-23 are correlated during the clinical course of dengue infection. A positive correlation was found between IL-17A and IL-23 levels (r = 0.4089, p =
    Figure Legend Snippet: Serum levels of IL-17A and IL-23 are correlated during the clinical course of dengue infection. A positive correlation was found between IL-17A and IL-23 levels (r = 0.4089, p =

    Techniques Used: Infection

    Dynamics of production of IL-17-related cytokines at different illness phases. Dengue patients were grouped according to illness phase (febrile, defervescence and convalescence) and infection type (primary and secondary). Median values of IL-17A serum levels are shown for primary ( A ) and secondary dengue infections ( B ), for IL-17F in primary ( C ) and secondary ( D ) infections, for IL-22 in primary ( E ) and secondary ( F ) infections and for IL-23 in primary ( G ) or secondary ( H ) infections. Individual data are shown and the horizontal line is the median value. Statistical difference is based on the Kruskal–Wallis with Dunn’s multiple comparison post-tests. * p
    Figure Legend Snippet: Dynamics of production of IL-17-related cytokines at different illness phases. Dengue patients were grouped according to illness phase (febrile, defervescence and convalescence) and infection type (primary and secondary). Median values of IL-17A serum levels are shown for primary ( A ) and secondary dengue infections ( B ), for IL-17F in primary ( C ) and secondary ( D ) infections, for IL-22 in primary ( E ) and secondary ( F ) infections and for IL-23 in primary ( G ) or secondary ( H ) infections. Individual data are shown and the horizontal line is the median value. Statistical difference is based on the Kruskal–Wallis with Dunn’s multiple comparison post-tests. * p

    Techniques Used: Infection

    IL-17A and IL-22 are associated with DHF severity. Serum levels of ( A ) IL-17A, ( B ) IL-17F, ( C ) IL-22 and ( D ) IL-23 in healthy controls (HC) ( n = 78) compared with dengue fever (DF, n = 97) and dengue hemorrhagic fever (DHF, n = 31). Individual data are shown and the horizontal line is the median value. Statistical difference is based on the Kruskal–Wallis with Dunn’s multiple comparison post-tests. * p
    Figure Legend Snippet: IL-17A and IL-22 are associated with DHF severity. Serum levels of ( A ) IL-17A, ( B ) IL-17F, ( C ) IL-22 and ( D ) IL-23 in healthy controls (HC) ( n = 78) compared with dengue fever (DF, n = 97) and dengue hemorrhagic fever (DHF, n = 31). Individual data are shown and the horizontal line is the median value. Statistical difference is based on the Kruskal–Wallis with Dunn’s multiple comparison post-tests. * p

    Techniques Used:

    Elevated serum levels of interleukin 17A (IL-17A), IL-17F, IL-22 and IL-23 are produced in dengue infections. Serum levels of ( A ) IL-17A, ( B ) IL-17F, ( C ) IL-22 and ( D ) IL-23 in healthy controls (HC) ( n = 78) compared with dengue virus (DENV)-infected patients ( n = 128). Individual data are shown and the horizontal line is the median value. Statistical difference is based on the Mann–Whitney test. ** p
    Figure Legend Snippet: Elevated serum levels of interleukin 17A (IL-17A), IL-17F, IL-22 and IL-23 are produced in dengue infections. Serum levels of ( A ) IL-17A, ( B ) IL-17F, ( C ) IL-22 and ( D ) IL-23 in healthy controls (HC) ( n = 78) compared with dengue virus (DENV)-infected patients ( n = 128). Individual data are shown and the horizontal line is the median value. Statistical difference is based on the Mann–Whitney test. ** p

    Techniques Used: Produced, Infection, MANN-WHITNEY

    11) Product Images from "Constitutive STAT3 phosphorylation and IL-6/IL-10 co-expression are associated with impaired T-cell function in tuberculosis patients"

    Article Title: Constitutive STAT3 phosphorylation and IL-6/IL-10 co-expression are associated with impaired T-cell function in tuberculosis patients

    Journal: Cellular and Molecular Immunology

    doi: 10.1038/cmi.2018.5

    M. tuberculosis PPD-specific and constitutive cytokine expression and plasma cytokine concentrations from tuberculosis patients and healthy contacts. Cytokine concentrations of IFN-γ, TNF-α, IL-17F, IL-22, IL-21, IL-6, and IL-10 of supernatants from diluted whole blood after 72 h in vitro culture with ( a and b ) or without ( c and d ) M. tuberculosis PPD are depicted. Association of differentially expressed cytokines are shown in ( b ) for PPD-specific IL-17F and IL-22, and ( d ) for constitutive IL-6 and IL-10 values. ( e ) Plasma concentrations of IL-10, IL-21, IL-23, IL-27 are shown for healthy contacts and tuberculosis patients. The dotted line indicates the detection limit for the respective cytokines. Symbols placed on this line indicate values below the detection limit. All samples were measured in duplicate, and mean values are indicated as open circles for healthy contacts and grey squares for tuberculosis patients. Study group medians and percentiles (25, 75) are shown. Significant differences are indicated by asterisks. Nominal P -values for the Mann-Whitney U -test (two-tailed) were calculated and shown as * for P
    Figure Legend Snippet: M. tuberculosis PPD-specific and constitutive cytokine expression and plasma cytokine concentrations from tuberculosis patients and healthy contacts. Cytokine concentrations of IFN-γ, TNF-α, IL-17F, IL-22, IL-21, IL-6, and IL-10 of supernatants from diluted whole blood after 72 h in vitro culture with ( a and b ) or without ( c and d ) M. tuberculosis PPD are depicted. Association of differentially expressed cytokines are shown in ( b ) for PPD-specific IL-17F and IL-22, and ( d ) for constitutive IL-6 and IL-10 values. ( e ) Plasma concentrations of IL-10, IL-21, IL-23, IL-27 are shown for healthy contacts and tuberculosis patients. The dotted line indicates the detection limit for the respective cytokines. Symbols placed on this line indicate values below the detection limit. All samples were measured in duplicate, and mean values are indicated as open circles for healthy contacts and grey squares for tuberculosis patients. Study group medians and percentiles (25, 75) are shown. Significant differences are indicated by asterisks. Nominal P -values for the Mann-Whitney U -test (two-tailed) were calculated and shown as * for P

    Techniques Used: Expressing, In Vitro, MANN-WHITNEY, Two Tailed Test

    12) Product Images from "Myeloid-Derived Suppressor Cells and γδT17 Cells Contribute to the Development of Gastric MALT Lymphoma in H. felis-Infected Mice"

    Article Title: Myeloid-Derived Suppressor Cells and γδT17 Cells Contribute to the Development of Gastric MALT Lymphoma in H. felis-Infected Mice

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2019.03104

    H. felis -infected mice show interleukin (IL)-1β activation and chemokine stimulation. (A) IL-1β and IL-23 levels in the gastric homogenates at 8 months postinfection. (B) IL-1β and IL-23 messenger RNA (mRNA) levels in the gastric tissue at 8, 11, 14, and 19 months postinfection. (C) TLR2, TLR7, and TLR9 mRNA levels at various time points after infection. (D) Immunoblot showing NLRP3 inflammasome expression in the stomach of H. felis -infected and control mice. (E) CCL20 and CCR6 mRNA levels in both groups. Data are shown as mean ± SD, * P
    Figure Legend Snippet: H. felis -infected mice show interleukin (IL)-1β activation and chemokine stimulation. (A) IL-1β and IL-23 levels in the gastric homogenates at 8 months postinfection. (B) IL-1β and IL-23 messenger RNA (mRNA) levels in the gastric tissue at 8, 11, 14, and 19 months postinfection. (C) TLR2, TLR7, and TLR9 mRNA levels at various time points after infection. (D) Immunoblot showing NLRP3 inflammasome expression in the stomach of H. felis -infected and control mice. (E) CCL20 and CCR6 mRNA levels in both groups. Data are shown as mean ± SD, * P

    Techniques Used: Infection, Mouse Assay, Activation Assay, Expressing

    The expansion of γδT17 cells and upregulation of interleukin (IL)-1β and IL-23 in mucosa-associated lymphoid tissue (MALT) lymphoma patients. (A) Representative immunofluorescence images showing γδT17 (TCRγδ + IL-17 + ) cells in gastric MALT lymphoma tissues and paired normal tissue. IL-17 in red, TCRγδ in green, and 4′ ,6-diamidino-2-phenylindole (DAPI) in blue; original magnification, 400×. (B) Percentage of γδT17 (TCRγδ + IL-17 + ) cells in the above groups. (C) Representative immunohistochemistry (IHC) images showing in situ IL-1β and IL-23 in gastric MALT lymphoma tissues and paired normal gastric tissues. (D) Histoscores of IL-1β and IL-23 were statistically higher in gastric MALT lymphoma tissues than control. Original magnification, 400×; ** P
    Figure Legend Snippet: The expansion of γδT17 cells and upregulation of interleukin (IL)-1β and IL-23 in mucosa-associated lymphoid tissue (MALT) lymphoma patients. (A) Representative immunofluorescence images showing γδT17 (TCRγδ + IL-17 + ) cells in gastric MALT lymphoma tissues and paired normal tissue. IL-17 in red, TCRγδ in green, and 4′ ,6-diamidino-2-phenylindole (DAPI) in blue; original magnification, 400×. (B) Percentage of γδT17 (TCRγδ + IL-17 + ) cells in the above groups. (C) Representative immunohistochemistry (IHC) images showing in situ IL-1β and IL-23 in gastric MALT lymphoma tissues and paired normal gastric tissues. (D) Histoscores of IL-1β and IL-23 were statistically higher in gastric MALT lymphoma tissues than control. Original magnification, 400×; ** P

    Techniques Used: Immunofluorescence, Immunohistochemistry, In Situ

    13) Product Images from "ICELLNET: a transcriptome-based framework to dissect intercellular communication"

    Article Title: ICELLNET: a transcriptome-based framework to dissect intercellular communication

    Journal: bioRxiv

    doi: 10.1101/2020.03.05.976878

    IL-10R blocking activates a cell-to-cell communication module in LPS-stimulated DCs. (A) Depicted are the 4 experimental conditions for which transcriptomics was generated (n = 6). (B) Connectivity maps describing outward communication from DCs to putative target cells in the conditions: Med, LPS, LPS+αTNFR and LPS+αIL-10R. Twelve primary cell types are considered as peripheral cells and are colored depending on the cell compartment (green: stroma, orange: innate, blue: adaptive, pink: epithelium). The width of the edges corresponds to a global score combining the intensity of all the individual ligand/receptor interactions, normalized to the medium condition. A scale ranging from 1 to 10, corresponding to minimum and maximum communication scores, is shown in the legend. (C) Gene corresponding to ligands (black) and receptors (white) counted in each loop signature and plotted according to regulation directionality: upregulated (Up) or downregulated (Down). Genes with separability score ≥ 4 were included in each condition’s signature. ( D) Protein levels of IL-6, OSM, IL-23 and IL-12p70 (means ± SEM), demonstrating increased secretion in LPS+αIL-10R DC supernatant.
    Figure Legend Snippet: IL-10R blocking activates a cell-to-cell communication module in LPS-stimulated DCs. (A) Depicted are the 4 experimental conditions for which transcriptomics was generated (n = 6). (B) Connectivity maps describing outward communication from DCs to putative target cells in the conditions: Med, LPS, LPS+αTNFR and LPS+αIL-10R. Twelve primary cell types are considered as peripheral cells and are colored depending on the cell compartment (green: stroma, orange: innate, blue: adaptive, pink: epithelium). The width of the edges corresponds to a global score combining the intensity of all the individual ligand/receptor interactions, normalized to the medium condition. A scale ranging from 1 to 10, corresponding to minimum and maximum communication scores, is shown in the legend. (C) Gene corresponding to ligands (black) and receptors (white) counted in each loop signature and plotted according to regulation directionality: upregulated (Up) or downregulated (Down). Genes with separability score ≥ 4 were included in each condition’s signature. ( D) Protein levels of IL-6, OSM, IL-23 and IL-12p70 (means ± SEM), demonstrating increased secretion in LPS+αIL-10R DC supernatant.

    Techniques Used: Blocking Assay, Generated

    14) Product Images from "Microbiota-induced tissue signals regulate ILC3-mediated antigen presentation"

    Article Title: Microbiota-induced tissue signals regulate ILC3-mediated antigen presentation

    Journal: Nature Communications

    doi: 10.1038/s41467-020-15612-2

    The microbiota and IL-23 negatively regulate Ag presentation by ILC3s. a CFSE-labeled OT-II tg CD4 + T cells were stimulated with 1 × 10 4 SPF or GF ILC3s ( Rag −/− background) in the presence of Ova peptide. n = 12 distinct samples of three independent experiments. ILC3s were sorted as depicted in Supplementary Fig. 2a . b MHC II expression of SI ILC3s from SPF ( n = 6 mice) and GF ( n = 8 mice) mice on Rag −/− background. NCR − ILC3s were gated as shown in Fig. 1 . Two independent experiments. c Relative expression of Il12b and Il23p19 in the terminal ileum of SPF ( n = 6 mice) and GF ( n = 5 mice) mice on Rag −/− background analyzed by qRT PCR. Two independent experiments. d MHC II expression of SI ILC3s from WT and Il23p19 −/− mice ( n = 10 mice). NCR − ILC3s were gated as shown in Fig. 1 . Five independent experiments. e Naive CFSE-labeled OT-II tg CD4 + T cells were cultured either with 5 × 10 4 SI ILC3s of WT ( n = 8 distinct samples) or Il23p19 −/− ( n = 10 distinct samples) mice in the presence of Ova peptide. Four independent experiments. ILC3s were sorted as depicted in Supplementary Fig. 2a . f Surface expression of MHC II and CD74 on ILC3s of Rag2 −/− mice cultured 7 days with or without IL-23 in addition to IL-2, IL-7, and SCF ( n = 5 distinct samples). NCR − ILC3s were gated as shown in Fig. 1 . Five independent experiments. g SI ILC3s were sort-purified from Rag2 −/− mice (Supplementary Fig. 2a ) and cultured 7 days with or without IL-23 in addition to IL-2, IL-7, and SCF. CFSE-labeled OT-II tg CD4 + T cells were stimulated with 5 × 10 4 cultured ILC3s in the presence of Ova peptide. n = 9 distinct samples of three independent experiments. Each symbol represents a sample and the bar graph represents the mean ± s.e.m. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001, calculated with mixed-effects models (two-sided) using lmerTest ( a , e , and g ) or two-tailed unpaired ( b , c Il12b , and d ) or two-tailed paired ( f ) Student’s t test or two-tailed Mann–Whitney test ( c Il23p19 ). Source data are provided as a Source Data File.
    Figure Legend Snippet: The microbiota and IL-23 negatively regulate Ag presentation by ILC3s. a CFSE-labeled OT-II tg CD4 + T cells were stimulated with 1 × 10 4 SPF or GF ILC3s ( Rag −/− background) in the presence of Ova peptide. n = 12 distinct samples of three independent experiments. ILC3s were sorted as depicted in Supplementary Fig. 2a . b MHC II expression of SI ILC3s from SPF ( n = 6 mice) and GF ( n = 8 mice) mice on Rag −/− background. NCR − ILC3s were gated as shown in Fig. 1 . Two independent experiments. c Relative expression of Il12b and Il23p19 in the terminal ileum of SPF ( n = 6 mice) and GF ( n = 5 mice) mice on Rag −/− background analyzed by qRT PCR. Two independent experiments. d MHC II expression of SI ILC3s from WT and Il23p19 −/− mice ( n = 10 mice). NCR − ILC3s were gated as shown in Fig. 1 . Five independent experiments. e Naive CFSE-labeled OT-II tg CD4 + T cells were cultured either with 5 × 10 4 SI ILC3s of WT ( n = 8 distinct samples) or Il23p19 −/− ( n = 10 distinct samples) mice in the presence of Ova peptide. Four independent experiments. ILC3s were sorted as depicted in Supplementary Fig. 2a . f Surface expression of MHC II and CD74 on ILC3s of Rag2 −/− mice cultured 7 days with or without IL-23 in addition to IL-2, IL-7, and SCF ( n = 5 distinct samples). NCR − ILC3s were gated as shown in Fig. 1 . Five independent experiments. g SI ILC3s were sort-purified from Rag2 −/− mice (Supplementary Fig. 2a ) and cultured 7 days with or without IL-23 in addition to IL-2, IL-7, and SCF. CFSE-labeled OT-II tg CD4 + T cells were stimulated with 5 × 10 4 cultured ILC3s in the presence of Ova peptide. n = 9 distinct samples of three independent experiments. Each symbol represents a sample and the bar graph represents the mean ± s.e.m. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001, calculated with mixed-effects models (two-sided) using lmerTest ( a , e , and g ) or two-tailed unpaired ( b , c Il12b , and d ) or two-tailed paired ( f ) Student’s t test or two-tailed Mann–Whitney test ( c Il23p19 ). Source data are provided as a Source Data File.

    Techniques Used: Labeling, Expressing, Mouse Assay, Quantitative RT-PCR, Cell Culture, Purification, Two Tailed Test, MANN-WHITNEY

    IL-23 reduces MHC II expression through activation of mTORC1 and STAT3. a Phosphorylation of S6 of SI ILC3s (sorted from Flt3L tg mice) after 48 h stimulation with or without IL-23 and 0, 10, or 100 nM Rapamycin (red = tubulin and green = p-S6). ILC3s were sorted as depicted in Supplementary Fig. 2a . Indicated is the relative phosphorylation of S6 (fluorescence signal for p-S6 normalized by fluorescence signal for tubulin). The indicated bar represents 3 mm. n = 6 distinct samples of five independent experiments. b Phosphorylation of STAT3(Tyr705) of sort-purified SI ILC3s (Supplementary Fig. 2a ) from Rag2 −/− mice after 20 min stimulation with or without IL-23. n = 4 distinct samples of four independent experiments. c MHC II expression of SI and SP ILC3s from Rptor fl/fl Rag2 −/− and Rptor ILC3−/− Rag2 −/− mice. NCR − ILC3s were gated as shown in Fig. 1 . n = 6 mice of two independent experiments. d MHC II expression of SI and SP ILC3s from Stat3 fl/fl ( n = 4 mice) and Vav1-Cre tg Stat3 fl/fl ( n = 7 mice) mice. NCR − ILC3s were gated as shown in Fig. 1 . Two independent experiments. Each symbol represents a sample and the bar graph represents the mean ± s.e.m. n.s. not significant; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001, calculated with one-way ANOVA (two-tailed) and Bonferroni’s multiple comparisons test ( a ) or two-tailed paired ( b ) or two-tailed unpaired ( c and d ) Student’s t test. Source data are provided as a Source Data File.
    Figure Legend Snippet: IL-23 reduces MHC II expression through activation of mTORC1 and STAT3. a Phosphorylation of S6 of SI ILC3s (sorted from Flt3L tg mice) after 48 h stimulation with or without IL-23 and 0, 10, or 100 nM Rapamycin (red = tubulin and green = p-S6). ILC3s were sorted as depicted in Supplementary Fig. 2a . Indicated is the relative phosphorylation of S6 (fluorescence signal for p-S6 normalized by fluorescence signal for tubulin). The indicated bar represents 3 mm. n = 6 distinct samples of five independent experiments. b Phosphorylation of STAT3(Tyr705) of sort-purified SI ILC3s (Supplementary Fig. 2a ) from Rag2 −/− mice after 20 min stimulation with or without IL-23. n = 4 distinct samples of four independent experiments. c MHC II expression of SI and SP ILC3s from Rptor fl/fl Rag2 −/− and Rptor ILC3−/− Rag2 −/− mice. NCR − ILC3s were gated as shown in Fig. 1 . n = 6 mice of two independent experiments. d MHC II expression of SI and SP ILC3s from Stat3 fl/fl ( n = 4 mice) and Vav1-Cre tg Stat3 fl/fl ( n = 7 mice) mice. NCR − ILC3s were gated as shown in Fig. 1 . Two independent experiments. Each symbol represents a sample and the bar graph represents the mean ± s.e.m. n.s. not significant; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001, calculated with one-way ANOVA (two-tailed) and Bonferroni’s multiple comparisons test ( a ) or two-tailed paired ( b ) or two-tailed unpaired ( c and d ) Student’s t test. Source data are provided as a Source Data File.

    Techniques Used: Expressing, Activation Assay, Mouse Assay, Fluorescence, Purification, Two Tailed Test

    15) Product Images from "Microbiota-induced TNF-like ligand 1A drives group 3 innate lymphoid cell-mediated barrier protection and intestinal T cell activation during colitis"

    Article Title: Microbiota-induced TNF-like ligand 1A drives group 3 innate lymphoid cell-mediated barrier protection and intestinal T cell activation during colitis

    Journal: Immunity

    doi: 10.1016/j.immuni.2018.10.014

    TL1A potently and selectively synergizes with IL-23 via MAPK to induce IL- 22 production in ILC3 in vitro and in vivo . ( A, B ) Sorted ILC3s from the intestinal lamina propria from WT, Tnfrsf25 +I- (called DR3 +/− ) or Tnfrsf25 −/− (called DR3 −/− ) mice were stimulated ex vivo with media (−), rIL-23 and/or rTL1A. Quantitative PCR (A) or intracellular cytokine staining for IL-22 (B) was performed after 18h stimulation. One of three independent experiments is shown. ( C ) IL-22 production by sorted ILC3 subsets was measured by intracellular flow cytometry after 18h stimulation. Brefeldin was added to the cultures 4h before intracellular cytokine staining. One of two independent experiments is shown. ( D ) Intracellular phosphoflow analysis was performed on sorted ILC3 for phosphorylation of p38-MAPK, ERK and IкBa 30 minute following stimulation. One of two independent experiments is shown. ( E ) Sorted ILC3s were stimulated for with rIL-23 and rTL1A in the presence of the indicated soluble inhibitors or control carrier DMSO (NF-кB inhibitor, NBD, white; TAK1 inhibitor, 5Z-7-Oxozeaenol, red; p38 MAPK inhibitor, SB203580, blue). IL-22 production was evaluated by ELISA. One of three independent experiments is shown. ( F ) IL-22 production by sorted ILC3s from WT or Il1r −/− mice after 18 hour stimulation with media, rIL-1β, rTL1A, or rIL-23 (or in combinations). ( G ) Survival curves of WT, DR3 −/− , Il23r GFP/GFP and Tnfrsf25 −/− Il23r GFP/GFP (called DR3 −/− I/23r GFP/GFP ) mice following 7 day treatment with 2% DSS. Data in are compiled from two independent experiments (with N=26, 18, 7, and 4 mice/group, respectively). Data in A, E, and F are mean ± SEM and analyzed by ANOVA (A, E) or two-tailed Student’s t-test with Bonferroni correction. Data in G are analyzed by Log-rank (Mantel-Cox) test; ** P
    Figure Legend Snippet: TL1A potently and selectively synergizes with IL-23 via MAPK to induce IL- 22 production in ILC3 in vitro and in vivo . ( A, B ) Sorted ILC3s from the intestinal lamina propria from WT, Tnfrsf25 +I- (called DR3 +/− ) or Tnfrsf25 −/− (called DR3 −/− ) mice were stimulated ex vivo with media (−), rIL-23 and/or rTL1A. Quantitative PCR (A) or intracellular cytokine staining for IL-22 (B) was performed after 18h stimulation. One of three independent experiments is shown. ( C ) IL-22 production by sorted ILC3 subsets was measured by intracellular flow cytometry after 18h stimulation. Brefeldin was added to the cultures 4h before intracellular cytokine staining. One of two independent experiments is shown. ( D ) Intracellular phosphoflow analysis was performed on sorted ILC3 for phosphorylation of p38-MAPK, ERK and IкBa 30 minute following stimulation. One of two independent experiments is shown. ( E ) Sorted ILC3s were stimulated for with rIL-23 and rTL1A in the presence of the indicated soluble inhibitors or control carrier DMSO (NF-кB inhibitor, NBD, white; TAK1 inhibitor, 5Z-7-Oxozeaenol, red; p38 MAPK inhibitor, SB203580, blue). IL-22 production was evaluated by ELISA. One of three independent experiments is shown. ( F ) IL-22 production by sorted ILC3s from WT or Il1r −/− mice after 18 hour stimulation with media, rIL-1β, rTL1A, or rIL-23 (or in combinations). ( G ) Survival curves of WT, DR3 −/− , Il23r GFP/GFP and Tnfrsf25 −/− Il23r GFP/GFP (called DR3 −/− I/23r GFP/GFP ) mice following 7 day treatment with 2% DSS. Data in are compiled from two independent experiments (with N=26, 18, 7, and 4 mice/group, respectively). Data in A, E, and F are mean ± SEM and analyzed by ANOVA (A, E) or two-tailed Student’s t-test with Bonferroni correction. Data in G are analyzed by Log-rank (Mantel-Cox) test; ** P

    Techniques Used: In Vitro, In Vivo, Mouse Assay, Ex Vivo, Real-time Polymerase Chain Reaction, Staining, Flow Cytometry, Cytometry, Enzyme-linked Immunosorbent Assay, Two Tailed Test

    16) Product Images from "GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis"

    Article Title: GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.01520

    GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p
    Figure Legend Snippet: GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p

    Techniques Used: Recombinant, Enzyme-linked Immunosorbent Assay

    17) Product Images from "Microbiota-induced TNF-like ligand 1A drives group 3 innate lymphoid cell-mediated barrier protection and intestinal T cell activation during colitis"

    Article Title: Microbiota-induced TNF-like ligand 1A drives group 3 innate lymphoid cell-mediated barrier protection and intestinal T cell activation during colitis

    Journal: Immunity

    doi: 10.1016/j.immuni.2018.10.014

    TL1A potently and selectively synergizes with IL-23 via MAPK to induce IL- 22 production in ILC3 in vitro and in vivo . ( A, B ) Sorted ILC3s from the intestinal lamina propria from WT, Tnfrsf25 +I- (called DR3 +/− ) or Tnfrsf25 −/− (called DR3 −/− ) mice were stimulated ex vivo with media (−), rIL-23 and/or rTL1A. Quantitative PCR (A) or intracellular cytokine staining for IL-22 (B) was performed after 18h stimulation. One of three independent experiments is shown. ( C ) IL-22 production by sorted ILC3 subsets was measured by intracellular flow cytometry after 18h stimulation. Brefeldin was added to the cultures 4h before intracellular cytokine staining. One of two independent experiments is shown. ( D ) Intracellular phosphoflow analysis was performed on sorted ILC3 for phosphorylation of p38-MAPK, ERK and IкBa 30 minute following stimulation. One of two independent experiments is shown. ( E ) Sorted ILC3s were stimulated for with rIL-23 and rTL1A in the presence of the indicated soluble inhibitors or control carrier DMSO (NF-кB inhibitor, NBD, white; TAK1 inhibitor, 5Z-7-Oxozeaenol, red; p38 MAPK inhibitor, SB203580, blue). IL-22 production was evaluated by ELISA. One of three independent experiments is shown. ( F ) IL-22 production by sorted ILC3s from WT or Il1r −/− mice after 18 hour stimulation with media, rIL-1β, rTL1A, or rIL-23 (or in combinations). ( G ) Survival curves of WT, DR3 −/− , Il23r GFP/GFP and Tnfrsf25 −/− Il23r GFP/GFP (called DR3 −/− I/23r GFP/GFP ) mice following 7 day treatment with 2% DSS. Data in are compiled from two independent experiments (with N=26, 18, 7, and 4 mice/group, respectively). Data in A, E, and F are mean ± SEM and analyzed by ANOVA (A, E) or two-tailed Student’s t-test with Bonferroni correction. Data in G are analyzed by Log-rank (Mantel-Cox) test; ** P
    Figure Legend Snippet: TL1A potently and selectively synergizes with IL-23 via MAPK to induce IL- 22 production in ILC3 in vitro and in vivo . ( A, B ) Sorted ILC3s from the intestinal lamina propria from WT, Tnfrsf25 +I- (called DR3 +/− ) or Tnfrsf25 −/− (called DR3 −/− ) mice were stimulated ex vivo with media (−), rIL-23 and/or rTL1A. Quantitative PCR (A) or intracellular cytokine staining for IL-22 (B) was performed after 18h stimulation. One of three independent experiments is shown. ( C ) IL-22 production by sorted ILC3 subsets was measured by intracellular flow cytometry after 18h stimulation. Brefeldin was added to the cultures 4h before intracellular cytokine staining. One of two independent experiments is shown. ( D ) Intracellular phosphoflow analysis was performed on sorted ILC3 for phosphorylation of p38-MAPK, ERK and IкBa 30 minute following stimulation. One of two independent experiments is shown. ( E ) Sorted ILC3s were stimulated for with rIL-23 and rTL1A in the presence of the indicated soluble inhibitors or control carrier DMSO (NF-кB inhibitor, NBD, white; TAK1 inhibitor, 5Z-7-Oxozeaenol, red; p38 MAPK inhibitor, SB203580, blue). IL-22 production was evaluated by ELISA. One of three independent experiments is shown. ( F ) IL-22 production by sorted ILC3s from WT or Il1r −/− mice after 18 hour stimulation with media, rIL-1β, rTL1A, or rIL-23 (or in combinations). ( G ) Survival curves of WT, DR3 −/− , Il23r GFP/GFP and Tnfrsf25 −/− Il23r GFP/GFP (called DR3 −/− I/23r GFP/GFP ) mice following 7 day treatment with 2% DSS. Data in are compiled from two independent experiments (with N=26, 18, 7, and 4 mice/group, respectively). Data in A, E, and F are mean ± SEM and analyzed by ANOVA (A, E) or two-tailed Student’s t-test with Bonferroni correction. Data in G are analyzed by Log-rank (Mantel-Cox) test; ** P

    Techniques Used: In Vitro, In Vivo, Mouse Assay, Ex Vivo, Real-time Polymerase Chain Reaction, Staining, Flow Cytometry, Cytometry, Enzyme-linked Immunosorbent Assay, Two Tailed Test

    18) Product Images from "Monocytic Cytokines in Autoimmune Polyglandular Syndrome Type 2 Are Modulated by Vitamin D and HLA-DQ"

    Article Title: Monocytic Cytokines in Autoimmune Polyglandular Syndrome Type 2 Are Modulated by Vitamin D and HLA-DQ

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.583709

    Vitamin D mediated reduction of pro-inflammatory cytokines. Gene expression (A, C, E, G) and protein secretion (B, D, F) of pro-inflammatory cytokines IL-6, CCL-2, IL-23, and IL-15 after in-vitro 1,25(OH) 2 D 3 in both APS-2 cohorts and HC. Data are presented as median values obtained from 15 AD/AIT patients, 15 T1D/AIT patients and 30 healthy controls (HC). T1D, type 1 diabetes; AIT, autoimmune thyroiditis; AD, Addison’s disease; HC, healthy controls. Values ​​are considered to be statistically significant when p
    Figure Legend Snippet: Vitamin D mediated reduction of pro-inflammatory cytokines. Gene expression (A, C, E, G) and protein secretion (B, D, F) of pro-inflammatory cytokines IL-6, CCL-2, IL-23, and IL-15 after in-vitro 1,25(OH) 2 D 3 in both APS-2 cohorts and HC. Data are presented as median values obtained from 15 AD/AIT patients, 15 T1D/AIT patients and 30 healthy controls (HC). T1D, type 1 diabetes; AIT, autoimmune thyroiditis; AD, Addison’s disease; HC, healthy controls. Values ​​are considered to be statistically significant when p

    Techniques Used: Expressing, In Vitro

    19) Product Images from "Oleanolic acid ameliorates intestinal alterations associated with EAE"

    Article Title: Oleanolic acid ameliorates intestinal alterations associated with EAE

    Journal: Journal of Neuroinflammation

    doi: 10.1186/s12974-020-02042-6

    OA treatment modulates inflammatory parameters in colon tissue from EAE mice. Levels in colon of a the inflammatory mediators TNFα, IL-1β, IL-23, IL-17, IGF-1, GM-SCF, and KC. b The Th2-type cytokine IL-23 and c the atypical cytokines IL-25 and IL-33 quantified by ELISAs. d Protein expression of NLRP6 and p-P65-NFκB in colon assessed by western blot. Bar graphs represent the mean ± SEM of 5–7 animals. * p
    Figure Legend Snippet: OA treatment modulates inflammatory parameters in colon tissue from EAE mice. Levels in colon of a the inflammatory mediators TNFα, IL-1β, IL-23, IL-17, IGF-1, GM-SCF, and KC. b The Th2-type cytokine IL-23 and c the atypical cytokines IL-25 and IL-33 quantified by ELISAs. d Protein expression of NLRP6 and p-P65-NFκB in colon assessed by western blot. Bar graphs represent the mean ± SEM of 5–7 animals. * p

    Techniques Used: Mouse Assay, Expressing, Western Blot

    20) Product Images from "Allogeneic ADSCs Induce the Production of Alloreactive Memory-CD8 T Cells through HLA-ABC Antigens"

    Article Title: Allogeneic ADSCs Induce the Production of Alloreactive Memory-CD8 T Cells through HLA-ABC Antigens

    Journal: Cells

    doi: 10.3390/cells9051246

    Effect of pro-inflammatory cytokines on the expression of human leukocyte antigens (HLAs) and co-stimulatory molecules on the surface of ADSCs. ADSCs were isolated from human adipose tissue and cultured in complete Dulbecco’s modified Eagle’s medium (DMEM). ADSCs that had been passaged fewer than eight times were used. ADSCs were stained with monoclonal antibodies (mAbs) against CD80 and CD86. ADSCs were additionally stained with mAbs against HLA-DR and HLA-ABC and for markers of NKG2DL with mAbs against MIC-A, MIC-B, ULBP1 and ULBP2/5/6. The data are representative of at least three experiments. IFN+IL-17: combination of IFN-γ and IL-17A/F; IFN+IL-17+IL-23: combination of IFN-γ, IL-17A/F and IL-23.
    Figure Legend Snippet: Effect of pro-inflammatory cytokines on the expression of human leukocyte antigens (HLAs) and co-stimulatory molecules on the surface of ADSCs. ADSCs were isolated from human adipose tissue and cultured in complete Dulbecco’s modified Eagle’s medium (DMEM). ADSCs that had been passaged fewer than eight times were used. ADSCs were stained with monoclonal antibodies (mAbs) against CD80 and CD86. ADSCs were additionally stained with mAbs against HLA-DR and HLA-ABC and for markers of NKG2DL with mAbs against MIC-A, MIC-B, ULBP1 and ULBP2/5/6. The data are representative of at least three experiments. IFN+IL-17: combination of IFN-γ and IL-17A/F; IFN+IL-17+IL-23: combination of IFN-γ, IL-17A/F and IL-23.

    Techniques Used: Expressing, Isolation, Cell Culture, Modification, Staining

    21) Product Images from "Phenotypic and functional markers for 1?,25-dihydroxyvitamin D3-modified regulatory dendritic cells"

    Article Title: Phenotypic and functional markers for 1?,25-dihydroxyvitamin D3-modified regulatory dendritic cells

    Journal:

    doi: 10.1111/j.1365-2249.2009.03961.x

    Cytokine secretion by 1α,25-dihydroxyvitamin D 3 -treated dendritic cells (VD3-DCs). (a) Secretion of interleukin (IL)-12p70, IL-10 and IL-23 by VD3-DCs: DCs were generated in the presence (‘VD3’) or absence [‘UT’
    Figure Legend Snippet: Cytokine secretion by 1α,25-dihydroxyvitamin D 3 -treated dendritic cells (VD3-DCs). (a) Secretion of interleukin (IL)-12p70, IL-10 and IL-23 by VD3-DCs: DCs were generated in the presence (‘VD3’) or absence [‘UT’

    Techniques Used: Generated

    22) Product Images from "Microbiota-dependent expansion of testicular IL-17-producing Vγ6+ γδ T cells upon puberty promotes local tissue immune surveillance"

    Article Title: Microbiota-dependent expansion of testicular IL-17-producing Vγ6+ γδ T cells upon puberty promotes local tissue immune surveillance

    Journal: Mucosal Immunology

    doi: 10.1038/s41385-020-0330-6

    Accumulation of γδ T cells in the testis is dependent on microbiota, IL-23 and TLR4 signalling. a Representative contour plots depicting IL-17-producing γδ T cells gated on CD3 + CD45 + cells in testes of specific pathogen-free (SPF) (left) and germ-free (GF) (right) mice. Scatter plot shows frequencies of γδ and γδ17 T cells among lymphocytes in testes of SPF (black) and GF (white) mice ( n = 18–22, five independent experiments). b Number of testicular γδ and γδ17 cells of SPF (black) and GF (white) mice ( n = 11–12, three independent experiments). c Representative contour plots depicting testicular Vγ6 + γδ T cells gated on CD3 + CD45 + cells of SPF and GF mice. Scatter plot displays frequencies of Vγ6 + γδ T cells among all T cells in testes of SPF (black) and GF (white) mice ( n = 11–14, four independent experiments). d Number of Vγ6 + γδ T cells from SPF and GF in pre- and post-pubertal mice ( n = 6, two independent experiments) ( n = 7–8, three independent experiments). e Scatter plot shows frequencies of γδ17 T cells among lymphocytes of WT (black), Tlr2 −/− , Tlr4 −/− and Myd88 −/− (white) mice ( n = 4–9, one to two independent experiments). f Representative histogram of γδ (dark grey) and αβ (white) T cells expressing IL-1 receptor (R) in WT mice (left) and IL-23R in Il23r gfp/gfp mice (right) and scatter plot with mean fluorescence intensity (MFI) ( n = 5–10, one to three independent experiments). g Scatter plot depicts frequencies of γδ17 among all T cells in testes of WT (black), Il1r −/− , Il23r −/− , Il1α −/− and Il1β − /− (white) ( n = 5–29, one to three independent experiments). h Scatter plot displays picogram (pg) per mg protein of IL-1α, IL-1β and IL-23 in the testis ( n = 9, two independent experiments). Data are represented as mean ± SD as evaluated by Kruskal–Wallis test followed by Dunn’s multiple-comparison test or one-way ANOVA followed by Holm–Sidak’s multiple-comparison test. ** P
    Figure Legend Snippet: Accumulation of γδ T cells in the testis is dependent on microbiota, IL-23 and TLR4 signalling. a Representative contour plots depicting IL-17-producing γδ T cells gated on CD3 + CD45 + cells in testes of specific pathogen-free (SPF) (left) and germ-free (GF) (right) mice. Scatter plot shows frequencies of γδ and γδ17 T cells among lymphocytes in testes of SPF (black) and GF (white) mice ( n = 18–22, five independent experiments). b Number of testicular γδ and γδ17 cells of SPF (black) and GF (white) mice ( n = 11–12, three independent experiments). c Representative contour plots depicting testicular Vγ6 + γδ T cells gated on CD3 + CD45 + cells of SPF and GF mice. Scatter plot displays frequencies of Vγ6 + γδ T cells among all T cells in testes of SPF (black) and GF (white) mice ( n = 11–14, four independent experiments). d Number of Vγ6 + γδ T cells from SPF and GF in pre- and post-pubertal mice ( n = 6, two independent experiments) ( n = 7–8, three independent experiments). e Scatter plot shows frequencies of γδ17 T cells among lymphocytes of WT (black), Tlr2 −/− , Tlr4 −/− and Myd88 −/− (white) mice ( n = 4–9, one to two independent experiments). f Representative histogram of γδ (dark grey) and αβ (white) T cells expressing IL-1 receptor (R) in WT mice (left) and IL-23R in Il23r gfp/gfp mice (right) and scatter plot with mean fluorescence intensity (MFI) ( n = 5–10, one to three independent experiments). g Scatter plot depicts frequencies of γδ17 among all T cells in testes of WT (black), Il1r −/− , Il23r −/− , Il1α −/− and Il1β − /− (white) ( n = 5–29, one to three independent experiments). h Scatter plot displays picogram (pg) per mg protein of IL-1α, IL-1β and IL-23 in the testis ( n = 9, two independent experiments). Data are represented as mean ± SD as evaluated by Kruskal–Wallis test followed by Dunn’s multiple-comparison test or one-way ANOVA followed by Holm–Sidak’s multiple-comparison test. ** P

    Techniques Used: Mouse Assay, Expressing, Fluorescence

    23) Product Images from "Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression"

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression

    Journal: The FASEB Journal

    doi: 10.1096/fj.201802433R

    IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    Figure Legend Snippet: IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    24) Product Images from "The immunomodulatory potential of the arylmethylaminosteroid sc1o"

    Article Title: The immunomodulatory potential of the arylmethylaminosteroid sc1o

    Journal: Journal of Molecular Medicine (Berlin, Germany)

    doi: 10.1007/s00109-020-02024-4

    Effect of sc1o on activated MdDCs. Human monocytes were differentiated to MdDCs for 5 days with GM-CSF (10 ng/ml) and IL-4 (10 ng/ml). MdDCs were activated with a mixture of cytokines (TNF-α, IL-6, IL-1β) and PGE2 in the presence or absence of different concentrations of sc1o (0.5–50 μM) or a vehicle (DMSO) for 24 h. ( a ) Surface marker expression was measured with a MACSQuant® Analyser 10 in triplicate. Fold induction of the geometric mean of the fluorescence intensity was calculated by referring treated cells to vehicle controls ( n = 6 different blood donors in 5 separate experiments). ( b ) Released concentrations of IL-23 in the supernatant were measured with ELISA in triplicate ( n = 6 different blood donors in 5 separate experiments). For statistical analysis, a one-way ANOVA with Dunnett’s multiple comparisons test ( a – b ) was used. Results are presented as means ± standard errors. * p
    Figure Legend Snippet: Effect of sc1o on activated MdDCs. Human monocytes were differentiated to MdDCs for 5 days with GM-CSF (10 ng/ml) and IL-4 (10 ng/ml). MdDCs were activated with a mixture of cytokines (TNF-α, IL-6, IL-1β) and PGE2 in the presence or absence of different concentrations of sc1o (0.5–50 μM) or a vehicle (DMSO) for 24 h. ( a ) Surface marker expression was measured with a MACSQuant® Analyser 10 in triplicate. Fold induction of the geometric mean of the fluorescence intensity was calculated by referring treated cells to vehicle controls ( n = 6 different blood donors in 5 separate experiments). ( b ) Released concentrations of IL-23 in the supernatant were measured with ELISA in triplicate ( n = 6 different blood donors in 5 separate experiments). For statistical analysis, a one-way ANOVA with Dunnett’s multiple comparisons test ( a – b ) was used. Results are presented as means ± standard errors. * p

    Techniques Used: Marker, Expressing, Fluorescence, Enzyme-linked Immunosorbent Assay

    25) Product Images from "Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression"

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression

    Journal: The FASEB Journal

    doi: 10.1096/fj.201802433R

    IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    Figure Legend Snippet: IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    26) Product Images from "GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis"

    Article Title: GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.01520

    GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p
    Figure Legend Snippet: GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p

    Techniques Used: Recombinant, Enzyme-linked Immunosorbent Assay

    27) Product Images from "Interleukin 17A controls Interleukin 17F production and maintains blood neutrophil counts in mice"

    Article Title: Interleukin 17A controls Interleukin 17F production and maintains blood neutrophil counts in mice

    Journal:

    doi: 10.4049/jimmunol.0804080

    Increased IL-17F response to IL-23 in Il17a -/- mice
    Figure Legend Snippet: Increased IL-17F response to IL-23 in Il17a -/- mice

    Techniques Used: Mouse Assay

    28) Product Images from "GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis"

    Article Title: GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.01520

    GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p
    Figure Legend Snippet: GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p

    Techniques Used: Recombinant, Enzyme-linked Immunosorbent Assay

    29) Product Images from "Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression"

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression

    Journal: The FASEB Journal

    doi: 10.1096/fj.201802433R

    IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    Figure Legend Snippet: IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    30) Product Images from "Combination Therapy of Acarbose and Cyclosporine a Ameliorates Imiquimod-Induced Psoriasis-Like Dermatitis in Mice"

    Article Title: Combination Therapy of Acarbose and Cyclosporine a Ameliorates Imiquimod-Induced Psoriasis-Like Dermatitis in Mice

    Journal: Molecules

    doi: 10.3390/molecules25081822

    Effect of Acar (250 mg/kg) and/or low-dose CsA (10 or 20 mg/kg) on serum and skin cytokines in mice with IMQ-induced psoriasis-like dermatitis on a daily basis for 9 days. The levels of interleukin (IL)-17A, IL-22, IL-23, and tumor necrosis factor (TNF)-α in the serum ( A ) and skin ( B ) were measured using ELISA. IMQ-induced mice were randomly assigned to 6 groups ( n = 6): IMQ alone, Acar (250 mg/kg/day), CsA (10 mg/kg/day), CsA (20 mg/kg/day), Acar (250 mg/kg/day) + CsA (10 mg/kg/day), and Acar (250 mg/kg/day) + CsA (20 mg/kg/day). The mice were subjected to an intervention with either Acar or CsA alone or in combination per day for 9 consecutive days orally. Experimental data are expressed as the mean ± SD ( n = 6) and data analysis was performed with Mann–Whitney U test. Columns with different superscript letters (a, b, ab) are significantly different at p
    Figure Legend Snippet: Effect of Acar (250 mg/kg) and/or low-dose CsA (10 or 20 mg/kg) on serum and skin cytokines in mice with IMQ-induced psoriasis-like dermatitis on a daily basis for 9 days. The levels of interleukin (IL)-17A, IL-22, IL-23, and tumor necrosis factor (TNF)-α in the serum ( A ) and skin ( B ) were measured using ELISA. IMQ-induced mice were randomly assigned to 6 groups ( n = 6): IMQ alone, Acar (250 mg/kg/day), CsA (10 mg/kg/day), CsA (20 mg/kg/day), Acar (250 mg/kg/day) + CsA (10 mg/kg/day), and Acar (250 mg/kg/day) + CsA (20 mg/kg/day). The mice were subjected to an intervention with either Acar or CsA alone or in combination per day for 9 consecutive days orally. Experimental data are expressed as the mean ± SD ( n = 6) and data analysis was performed with Mann–Whitney U test. Columns with different superscript letters (a, b, ab) are significantly different at p

    Techniques Used: Mouse Assay, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

    31) Product Images from "Elucidating the Role of Ezh2 in Tolerogenic Function of NOD Bone Marrow-Derived Dendritic Cells Expressing Constitutively Active Stat5b"

    Article Title: Elucidating the Role of Ezh2 in Tolerogenic Function of NOD Bone Marrow-Derived Dendritic Cells Expressing Constitutively Active Stat5b

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms21186453

    Cytokine profiles of BMDCs derived from NOD and NOD.CD11c Stat5b-CA mice. BMDCs (1 × 10 5 cells/well) generated from NOD and NOD.CD11c Stat5b-CA mice were cultured for 48 h in the absence or presence of GM-CSF (50 ng/mL). ( A ) Relative expression of Il12a , Il12b , Il23a , Il27a , and Tgfβ genes was determined by qPCR using the ΔΔCT method. qPCR data are shown as relative expression compared to untreated BMDCs of NOD mice. ( B ) Quantification of TGFβ, TNFα, and IL-23 released in the supernatants of BMDCs exposed or not to GM-CSF as determined by ELISA. Data are shown as the mean ± SEM of at least three independent experiments. The asterisks indicate statistically significant differences determined using one-way ANOVA with Tukey’s post-test. p
    Figure Legend Snippet: Cytokine profiles of BMDCs derived from NOD and NOD.CD11c Stat5b-CA mice. BMDCs (1 × 10 5 cells/well) generated from NOD and NOD.CD11c Stat5b-CA mice were cultured for 48 h in the absence or presence of GM-CSF (50 ng/mL). ( A ) Relative expression of Il12a , Il12b , Il23a , Il27a , and Tgfβ genes was determined by qPCR using the ΔΔCT method. qPCR data are shown as relative expression compared to untreated BMDCs of NOD mice. ( B ) Quantification of TGFβ, TNFα, and IL-23 released in the supernatants of BMDCs exposed or not to GM-CSF as determined by ELISA. Data are shown as the mean ± SEM of at least three independent experiments. The asterisks indicate statistically significant differences determined using one-way ANOVA with Tukey’s post-test. p

    Techniques Used: Derivative Assay, Mouse Assay, Generated, Cell Culture, Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    Inhibition of Ezh2 in Stat5b-CA-expressing BMDCs disturbs their tolerogenic signature and their capacity to protect against diabetes in NOD mice. ( A , B ) Stat5b-CA.BMDCs were pre-incubated for 1 h with the Ezh2 inhibitor GSK343 (3 µM) or vehicle (0.1% DMSO), then cultured for 48 h in the absence or presence of GM-CSF (50 ng/mL) and analyzed for ( A ) the expression of CD80, CD86, PD-L1, and PD-L2 by FACS and ( B ) production of cytokines (TGFβ, IL-10, IL-12, and IL-23) by ELISA. ( C ) BMDCs derived from NOD or NOD.CD11c Stat5b-CA mice were cultured in vitro with Ezh2 inhibitor GSK343 (3 µM) or vehicle (0.1% DMSO). DCs were washed and i.v. injected (10 7 cells/mouse) to 8-week-old female NOD mice (6 mice per group). Recipient NOD mice were followed for diabetes development until 26 weeks (22 weeks post-injection) of age. The asterisks indicate statistically significant differences determined by one-way ANOVA with Tukey’s post-test. p
    Figure Legend Snippet: Inhibition of Ezh2 in Stat5b-CA-expressing BMDCs disturbs their tolerogenic signature and their capacity to protect against diabetes in NOD mice. ( A , B ) Stat5b-CA.BMDCs were pre-incubated for 1 h with the Ezh2 inhibitor GSK343 (3 µM) or vehicle (0.1% DMSO), then cultured for 48 h in the absence or presence of GM-CSF (50 ng/mL) and analyzed for ( A ) the expression of CD80, CD86, PD-L1, and PD-L2 by FACS and ( B ) production of cytokines (TGFβ, IL-10, IL-12, and IL-23) by ELISA. ( C ) BMDCs derived from NOD or NOD.CD11c Stat5b-CA mice were cultured in vitro with Ezh2 inhibitor GSK343 (3 µM) or vehicle (0.1% DMSO). DCs were washed and i.v. injected (10 7 cells/mouse) to 8-week-old female NOD mice (6 mice per group). Recipient NOD mice were followed for diabetes development until 26 weeks (22 weeks post-injection) of age. The asterisks indicate statistically significant differences determined by one-way ANOVA with Tukey’s post-test. p

    Techniques Used: Inhibition, Expressing, Mouse Assay, Incubation, Cell Culture, FACS, Enzyme-linked Immunosorbent Assay, Derivative Assay, In Vitro, Injection

    32) Product Images from "GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis"

    Article Title: GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.01520

    GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p
    Figure Legend Snippet: GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p

    Techniques Used: Recombinant, Enzyme-linked Immunosorbent Assay

    33) Product Images from "IL-12 p40 monomer is different from other IL-12 family members to selectively inhibit IL-12Rβ1 internalization and suppress EAE"

    Article Title: IL-12 p40 monomer is different from other IL-12 family members to selectively inhibit IL-12Rβ1 internalization and suppress EAE

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.2000653117

    The p40 treatment retains the surface expression of IL-12Rβ1, but neither IL-12Rβ2 nor IL-23R, in MB P-primed T cells. ( A ) Splenocytes isolated from MBP-immunized donor mice were restimulated with MBP in the presence or absence of p40, IL-12, p40+IL-12, p40 2 , p40 2 +p40, IL-23, and IL-23+p40 for 4 h followed by FACS analysis of nonadherent cells in an LSRFortessa analyzer (BD Biosciences) for IL-12Rβ1 and CD4. Where cells were treated with the combination of cytokines, p40 was used 30 min prior to p40 2 , IL-12, or IL-23. ( B ) Under similar treatment conditions (p40, IL-12, p40+IL-12), the surface expression of IL-12Rβ2 was monitored by FACS. ( C ) Under similar treatment conditions (p40, IL-23, p40+IL-23), the surface expression of IL-23 was monitored by FACS. The MFI of IL-12Rβ1 ( D ), IL-12Rβ2 ( E ), and IL-23R ( F ) in the CD4 + population was calculated by using the CellQuest software. Data are mean ± SD of four different experiments. *** P
    Figure Legend Snippet: The p40 treatment retains the surface expression of IL-12Rβ1, but neither IL-12Rβ2 nor IL-23R, in MB P-primed T cells. ( A ) Splenocytes isolated from MBP-immunized donor mice were restimulated with MBP in the presence or absence of p40, IL-12, p40+IL-12, p40 2 , p40 2 +p40, IL-23, and IL-23+p40 for 4 h followed by FACS analysis of nonadherent cells in an LSRFortessa analyzer (BD Biosciences) for IL-12Rβ1 and CD4. Where cells were treated with the combination of cytokines, p40 was used 30 min prior to p40 2 , IL-12, or IL-23. ( B ) Under similar treatment conditions (p40, IL-12, p40+IL-12), the surface expression of IL-12Rβ2 was monitored by FACS. ( C ) Under similar treatment conditions (p40, IL-23, p40+IL-23), the surface expression of IL-23 was monitored by FACS. The MFI of IL-12Rβ1 ( D ), IL-12Rβ2 ( E ), and IL-23R ( F ) in the CD4 + population was calculated by using the CellQuest software. Data are mean ± SD of four different experiments. *** P

    Techniques Used: Expressing, Isolation, Mouse Assay, FACS, Software

    Enrichment of Tregs by p40. Splenocytes isolated from MBP-immunized donor mice were restimulated with MBP in the presence or absence of p40 (10 ng/mL), IL-23 (10 ng/mL), and p40+IL-23 for 48 h, followed by FACS analysis of nonadherent cells in an LSRFortessa analyzer (BD Biosciences) for CD4 and Foxp3 ( A ). The MFI of Foxp3 ( B ) in the CD4 + population was calculated by using the CellQuest software. Results are mean ± SD of three different experiments. **P
    Figure Legend Snippet: Enrichment of Tregs by p40. Splenocytes isolated from MBP-immunized donor mice were restimulated with MBP in the presence or absence of p40 (10 ng/mL), IL-23 (10 ng/mL), and p40+IL-23 for 48 h, followed by FACS analysis of nonadherent cells in an LSRFortessa analyzer (BD Biosciences) for CD4 and Foxp3 ( A ). The MFI of Foxp3 ( B ) in the CD4 + population was calculated by using the CellQuest software. Results are mean ± SD of three different experiments. **P

    Techniques Used: Isolation, Mouse Assay, FACS, Software

    34) Product Images from "Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression"

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression

    Journal: The FASEB Journal

    doi: 10.1096/fj.201802433R

    IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    Figure Legend Snippet: IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    35) Product Images from "Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression"

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression

    Journal: The FASEB Journal

    doi: 10.1096/fj.201802433R

    IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    Figure Legend Snippet: IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    36) Product Images from "Interleukin 17A controls Interleukin 17F production and maintains blood neutrophil counts in mice"

    Article Title: Interleukin 17A controls Interleukin 17F production and maintains blood neutrophil counts in mice

    Journal:

    doi: 10.4049/jimmunol.0804080

    Increased IL-17F response to IL-23 in Il17a -/- mice
    Figure Legend Snippet: Increased IL-17F response to IL-23 in Il17a -/- mice

    Techniques Used: Mouse Assay

    37) Product Images from "GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis"

    Article Title: GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.01520

    GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p
    Figure Legend Snippet: GM-CSF potentiates TNF and IL-23 production by LPS-stimulated whole blood from AS patients and healthy controls. Fresh whole blood from AS patients or healthy controls was treated with 10 ng/ml recombinant GM-CSF (A,B) , anti-GM-CSF or IgG control for 2 h (C,D) , then stimulated with 10 ng/ml LPS overnight. TNF (A,C) or IL-23 (B,D) measured in supernatants using ELISA. Data are represented as mean and SEM of independent donors in independent experiments. p -value was calculated using paired t- test. * p

    Techniques Used: Recombinant, Enzyme-linked Immunosorbent Assay

    38) Product Images from "Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression"

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression

    Journal: The FASEB Journal

    doi: 10.1096/fj.201802433R

    IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P
    Figure Legend Snippet: IL-1β and IL-23 promote IL-17A production from γδ T cells in the absence of TCR engagement. A ) Spleen cells from WT or Mif −/− BALB/c mice were stimulated with 20 ng/ml of IL-1β, 20 ng/ml of IL-23, or both for 72 h and stained for intracellular IL-17A and for surface CD3, CD4, and γδ TCRs. Numbers refer to percentage of CD3 + CD4 + IL-17A + cells or CD3 + γδTCR + IL-17A + cells. B ) LM induces IL-1β and IL-23 production by macrophages. BMDMs from WT or Mif −/− BALB/c mice were stimulated with 1 µg/ml of LM for 18 h. IL-1β and IL-23 concentrations in the supernatants were quantified by ELISA. Comp, compensation; ctrl, control; FL2-H, flourescence 2 channel height; Gr, granulocyte. Data shown are representative of 3 replicates with n = 4–5 per group. * P

    Techniques Used: Mouse Assay, Staining, Enzyme-linked Immunosorbent Assay

    39) Product Images from "Human Adipose Tissue-Derived Mesenchymal Stem Cells in Parkinson’s Disease: Inhibition of T Helper 17 Cell Differentiation and Regulation of Immune Balance Towards a Regulatory T Cell Phenotype"

    Article Title: Human Adipose Tissue-Derived Mesenchymal Stem Cells in Parkinson’s Disease: Inhibition of T Helper 17 Cell Differentiation and Regulation of Immune Balance Towards a Regulatory T Cell Phenotype

    Journal: Clinical Interventions in Aging

    doi: 10.2147/CIA.S259762

    Ad-MSCs suppressed the expression of cytokines and their receptors. Low levels of expression of IL-6R and IL-23R mRNAs in co-cultures under the Th17 polarized condition ( A, B ), and low levels of IL-6R mRNA under the Treg polarized condition ( C , n=3). Levels of IL-6, IL-23, and TGF-β proteins showed similar results in CD4 + T cell co-cultures under either of the polarized conditions ( D–F ). * p
    Figure Legend Snippet: Ad-MSCs suppressed the expression of cytokines and their receptors. Low levels of expression of IL-6R and IL-23R mRNAs in co-cultures under the Th17 polarized condition ( A, B ), and low levels of IL-6R mRNA under the Treg polarized condition ( C , n=3). Levels of IL-6, IL-23, and TGF-β proteins showed similar results in CD4 + T cell co-cultures under either of the polarized conditions ( D–F ). * p

    Techniques Used: Expressing

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    Article Title: Myeloid-Derived Suppressor Cells and γδT17 Cells Contribute to the Development of Gastric MALT Lymphoma in H. felis-Infected Mice
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    Cytokine Assay:

    Article Title: CD163+ cytokine-producing cDC2 stimulate intratumoral type 1 T cell responses in HPV16-induced oropharyngeal cancer
    Article Snippet: Cytokine production by CD163+ and CD163− myeloid cells CD11c+ HLA-DR+ and CD14+ CD163‒ , CD14+ CD163+ , CD14‒ CD163‒ and CD14‒ CD163+ myeloid cells were FACS-sorted from CD3‒ CD19‒ PBMCs from healthy donors and cultured in Iscove's Modified Dulbecco's Media (IMDM; Lonza) supplemented with 2% human AB (hAB) serum (Capricorn Scientific), 100 IU/mL penicillin (pen; Thermo Fisher Scientific), 100 µg/mL streptomycin (strep; Thermo Fisher Scientific) and 2 mmol/L l-glutamin (L-glut; Lonza) at 20,000 cells per well in triplo in a 96-well round bottom plate in the presence of 100 ng/mL lipopolysaccharide (LPS) (Sigma-Aldrich), 25 µg/mL poly(I:C, Invivogen) and 2.5 µg/mL R848 (Enzo life sciences). .. Cytokines were determined using IL-12p70 and IL-18 ELISA (R & D systems) and a custom-made 13-plex cytokine assay (Granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-1α, IL-1β, IL-10, IL-23, IL-6, IL-8, MIP-1α, MIP-3α, TGF-α, TNF-α, M-CSF and VEGF-A, ProcartaPlex Immunoassay, Thermo Fisher Scientific). .. Assessment of T cell stimulatory capacity of CD163+ and CD163− myeloid cells To assess the T cell stimulatory capacity of the flow cytometry-sorted CD163+ and CD163‒ myeloid subsets, these cells were cultured for 20 hours at 15.000–20.000 cells per well in triplo in IMDM + pen/strep/L-glut+2% hAB serum a 96-wells round bottom plate in the presence of 100 ng/mL LPS and 2.5 µg/mL R848, after which allogeneic Cell Trace Violet (CTV; Thermo Fisher Scientific)-labeled allogeneic PBMCs were added at 1:10, 1:20 and 1:40 myeloid-to-PBMC ratio.

    other:

    Article Title: GM-CSF Primes Proinflammatory Monocyte Responses in Ankylosing Spondylitis
    Article Snippet: Addition of GM-CSF significantly increased the production of TNF and IL-23 by LPS-stimulated whole blood from both AS patients and HC ( ).

    Produced:

    Article Title: Macrophage migration inhibitory factor regulates innate γδ T-cell responses via IL-17 expression
    Article Snippet: These results are consistent with a potential role for the up-regulated expression of IL-1R1 and IL-23R in the Mif -dependent expansion of γδ17 cells. .. IL-1β and IL-23 typically are produced by the innate response to pathogen-associated molecular patterns. .. Because γδ17 cells are activated during Mycobacterium infection , we tested whether the mycobacterial glycolipid LM stimulated IL-1β or IL-23 expression in Mif−/− macrophages.

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  • 93
    Thermo Fisher mouse interleukin 2 elisa test kit
    Direct DNA modification and capture of primary cells. (a) Human red blood cells were bound in the same manner as Jurkat cells on a DNA spot, and appeared to be morphologically identical immediately after binding. Trypan blue staining indicated that the membranes remained intact. The scale bar represents 25 µm. (b) DNA-coated mouse CD4 + helper T-cells were bound by spots coated with complementary DNA. After 3 min of exposure, a clear boundary can be seen between the printed and unprinted regions of the slide. (c) Microscale DNA patterns made by photolithography and microfabrication. Fluorescein conjugated ssDNA strands were patterned on the substrate to allow visualization. (d) Mouse primary T-cells were captured on the same DNA patterns. (e) <t>IL-2</t> production of DNA-immobilized T-cells and free T-cells, as determined by <t>ELISA.</t> ConA = concanavalin A; PMA = phorbolmeristyl acetate;CSA = cyclosporin A. In (b) and (d), the scale bars represent 100 µm.
    Mouse Interleukin 2 Elisa Test Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse interleukin 2 elisa test kit/product/Thermo Fisher
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    94
    Thermo Fisher il 2 human uncoated elisa kit
    Knockdown of GFAT1 or DON treatment in cancer cells increases T cell activation and NK cell killing. ( A ) Cancer cells were seeded in triplicate in 24-well plates at a density of 5 × 10 4 cells/well. The next day, cells were transfected with control or GFAT1 siRNA for another 24 h. The cells were then incubated with media containing IFNγ (10 ng/ml) for 6 h. The medium was then removed and Jurkat T cells in medium with 25 ng/ml PMA and 200 ug/ml ionomycin were added to cancer cells. The ratio of cancer cell to T cell was 1:2 for H2009 and 1:4 for SK-MES-1. The next day (18 h for H2009 and 24 h for SK-MES-1), the media were collected for <t>ELISA</t> for <t>IL-2</t> concentration. For DON inhibition, 10 μM DON was added overnight before the treatment of IFNγ, the ratio of cancer cell to T cell was 1:3 and incubation time was 24 h for both cell lines ( B ). ( C ) Cancer cells were seeded and transfected with siRNAs and treated with IFNγ as described in (A). NK cells were added to cancer cells in the indicated ratio for 4 h. The media were refreshed and NK cells removed. MTT assay was carried out 24 h later. *, P
    Il 2 Human Uncoated Elisa Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Thermo Fisher standard il 2 elisa kit
    Restoration of <t>IL-2</t> production in murine T REG by ectopic expression of NFAT2 Purified CD4 + CD25 neg Foxp3 neg T cells (first panel on the left in A and B ) and CD4 + CD25 + Foxp3 + T REG (right three panels in A and B ) were mock transduced (first and second panels in A and B ), transduced with empty MIGR1 vector (third panel in A and B ), or transduced with MIGR1 encoding murine NFAT2-IRES-GFP (last panel on right) and rested cultures were stained for Foxp3 ( A ) and CD25 (not shown). Transduced cells were cultured in medium (top panels in B ), or restimulated with plate-bound CD3/28 Ab (middle panels in B and in part C ) or PMA and ionomycin (bottom panels in B ), and IL-2 production was assessed by intracellular staining as a function of GFP reporter gene expression ( B ) or by <t>ELISA</t> ( C ). In a separate approach, murine CD4 + CD25 neg T cells were transduced with Foxp3-IRES-GFP vector alone (x-axis of left panel in D) , NFAT2-IRES-NGFR alone (y-axis of left panel in D ), or Foxp3-IRES-GFP plus NFAT2-IRES-NGFR in combination (right panel in D ). In the left panel, singly transduced cells were mixed prior to flow cytometry to demonstrate that each population of cells could be spectrally separated. The transduced cells depicted in D were restimulated with plate-bound anti-CD3/28 Ab, and IL-2 production was measured by intracellular staining with an APC-conjugated Ab. The graph in E shows the mean fluorescence intensity (MFI) of IL-2 staining as a function of GFP (NFAT2) or NGFR (Empty vector, Foxp3, and Foxp3 + NFAT2) reporter MFI. Results are representative of 3 experiments.
    Standard Il 2 Elisa Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    Thermo Fisher gene exp il2ra mm01340213 m1
    CD4+ <t>CD25+</t> FoxP3+ classical regulatory T cells subset in not increased by DCs after efferocytosis. Top: Percentage of proliferating CFSE low , CD3 + , CD4 + <t>,CD25</t> + , FoxP3 + regulatory T cells in autologous T cell proliferation assays with immature DCs (iDCs, white circles), mature DCs (mDCs, white squares) and DCs loaded with apoptotic cells (NITApo-DCs, black circles) with insulin (20 µg/ml) at a ratio of 1∶10 for 7 days. Plots show the mean (line) of four independent experiments. Double-sided Wilcoxon test was used for the evaluation of statistical significance. Bottom: Representative FACS plots showing CD25 + FoxP3 + regulatory T cells gated on CFSE low , CD3 + , CD4 + .
    Gene Exp Il2ra Mm01340213 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Direct DNA modification and capture of primary cells. (a) Human red blood cells were bound in the same manner as Jurkat cells on a DNA spot, and appeared to be morphologically identical immediately after binding. Trypan blue staining indicated that the membranes remained intact. The scale bar represents 25 µm. (b) DNA-coated mouse CD4 + helper T-cells were bound by spots coated with complementary DNA. After 3 min of exposure, a clear boundary can be seen between the printed and unprinted regions of the slide. (c) Microscale DNA patterns made by photolithography and microfabrication. Fluorescein conjugated ssDNA strands were patterned on the substrate to allow visualization. (d) Mouse primary T-cells were captured on the same DNA patterns. (e) IL-2 production of DNA-immobilized T-cells and free T-cells, as determined by ELISA. ConA = concanavalin A; PMA = phorbolmeristyl acetate;CSA = cyclosporin A. In (b) and (d), the scale bars represent 100 µm.

    Journal: Langmuir : the ACS journal of surfaces and colloids

    Article Title: Direct Cell Surface Modification with DNA for the Capture of Primary Cells and the Investigation of Myotube Formation on Defined Patterns

    doi: 10.1021/la900150n

    Figure Lengend Snippet: Direct DNA modification and capture of primary cells. (a) Human red blood cells were bound in the same manner as Jurkat cells on a DNA spot, and appeared to be morphologically identical immediately after binding. Trypan blue staining indicated that the membranes remained intact. The scale bar represents 25 µm. (b) DNA-coated mouse CD4 + helper T-cells were bound by spots coated with complementary DNA. After 3 min of exposure, a clear boundary can be seen between the printed and unprinted regions of the slide. (c) Microscale DNA patterns made by photolithography and microfabrication. Fluorescein conjugated ssDNA strands were patterned on the substrate to allow visualization. (d) Mouse primary T-cells were captured on the same DNA patterns. (e) IL-2 production of DNA-immobilized T-cells and free T-cells, as determined by ELISA. ConA = concanavalin A; PMA = phorbolmeristyl acetate;CSA = cyclosporin A. In (b) and (d), the scale bars represent 100 µm.

    Article Snippet: Portions of the culture media (1 mL) were withdrawn from each population of cells and tested for IL-2 production using a Mouse Interleukin-2 ELISA test kit (Thermo Scientific).

    Techniques: Modification, Binding Assay, Staining, Enzyme-linked Immunosorbent Assay

    Knockdown of GFAT1 or DON treatment in cancer cells increases T cell activation and NK cell killing. ( A ) Cancer cells were seeded in triplicate in 24-well plates at a density of 5 × 10 4 cells/well. The next day, cells were transfected with control or GFAT1 siRNA for another 24 h. The cells were then incubated with media containing IFNγ (10 ng/ml) for 6 h. The medium was then removed and Jurkat T cells in medium with 25 ng/ml PMA and 200 ug/ml ionomycin were added to cancer cells. The ratio of cancer cell to T cell was 1:2 for H2009 and 1:4 for SK-MES-1. The next day (18 h for H2009 and 24 h for SK-MES-1), the media were collected for ELISA for IL-2 concentration. For DON inhibition, 10 μM DON was added overnight before the treatment of IFNγ, the ratio of cancer cell to T cell was 1:3 and incubation time was 24 h for both cell lines ( B ). ( C ) Cancer cells were seeded and transfected with siRNAs and treated with IFNγ as described in (A). NK cells were added to cancer cells in the indicated ratio for 4 h. The media were refreshed and NK cells removed. MTT assay was carried out 24 h later. *, P

    Journal: bioRxiv

    Article Title: Inhibition of GFAT1 in lung cancer cells destabilizes PD-L1 protein and improves immune response

    doi: 10.1101/2020.11.17.385039

    Figure Lengend Snippet: Knockdown of GFAT1 or DON treatment in cancer cells increases T cell activation and NK cell killing. ( A ) Cancer cells were seeded in triplicate in 24-well plates at a density of 5 × 10 4 cells/well. The next day, cells were transfected with control or GFAT1 siRNA for another 24 h. The cells were then incubated with media containing IFNγ (10 ng/ml) for 6 h. The medium was then removed and Jurkat T cells in medium with 25 ng/ml PMA and 200 ug/ml ionomycin were added to cancer cells. The ratio of cancer cell to T cell was 1:2 for H2009 and 1:4 for SK-MES-1. The next day (18 h for H2009 and 24 h for SK-MES-1), the media were collected for ELISA for IL-2 concentration. For DON inhibition, 10 μM DON was added overnight before the treatment of IFNγ, the ratio of cancer cell to T cell was 1:3 and incubation time was 24 h for both cell lines ( B ). ( C ) Cancer cells were seeded and transfected with siRNAs and treated with IFNγ as described in (A). NK cells were added to cancer cells in the indicated ratio for 4 h. The media were refreshed and NK cells removed. MTT assay was carried out 24 h later. *, P

    Article Snippet: IL-2 concentration in culture medium released by Jurkat T cells was determined by an ELISA kit from Thermo-Fisher (Cat. 88-7025-88) according to the instruction of the manufacturer.

    Techniques: Activation Assay, Transfection, Incubation, Enzyme-linked Immunosorbent Assay, Concentration Assay, Inhibition, MTT Assay

    Restoration of IL-2 production in murine T REG by ectopic expression of NFAT2 Purified CD4 + CD25 neg Foxp3 neg T cells (first panel on the left in A and B ) and CD4 + CD25 + Foxp3 + T REG (right three panels in A and B ) were mock transduced (first and second panels in A and B ), transduced with empty MIGR1 vector (third panel in A and B ), or transduced with MIGR1 encoding murine NFAT2-IRES-GFP (last panel on right) and rested cultures were stained for Foxp3 ( A ) and CD25 (not shown). Transduced cells were cultured in medium (top panels in B ), or restimulated with plate-bound CD3/28 Ab (middle panels in B and in part C ) or PMA and ionomycin (bottom panels in B ), and IL-2 production was assessed by intracellular staining as a function of GFP reporter gene expression ( B ) or by ELISA ( C ). In a separate approach, murine CD4 + CD25 neg T cells were transduced with Foxp3-IRES-GFP vector alone (x-axis of left panel in D) , NFAT2-IRES-NGFR alone (y-axis of left panel in D ), or Foxp3-IRES-GFP plus NFAT2-IRES-NGFR in combination (right panel in D ). In the left panel, singly transduced cells were mixed prior to flow cytometry to demonstrate that each population of cells could be spectrally separated. The transduced cells depicted in D were restimulated with plate-bound anti-CD3/28 Ab, and IL-2 production was measured by intracellular staining with an APC-conjugated Ab. The graph in E shows the mean fluorescence intensity (MFI) of IL-2 staining as a function of GFP (NFAT2) or NGFR (Empty vector, Foxp3, and Foxp3 + NFAT2) reporter MFI. Results are representative of 3 experiments.

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: FOXP3 Inhibits Activation-Induced NFAT2 Expression in T Cells Thereby Limiting Effector Cytokine Expression

    doi: 10.4049/jimmunol.0800216

    Figure Lengend Snippet: Restoration of IL-2 production in murine T REG by ectopic expression of NFAT2 Purified CD4 + CD25 neg Foxp3 neg T cells (first panel on the left in A and B ) and CD4 + CD25 + Foxp3 + T REG (right three panels in A and B ) were mock transduced (first and second panels in A and B ), transduced with empty MIGR1 vector (third panel in A and B ), or transduced with MIGR1 encoding murine NFAT2-IRES-GFP (last panel on right) and rested cultures were stained for Foxp3 ( A ) and CD25 (not shown). Transduced cells were cultured in medium (top panels in B ), or restimulated with plate-bound CD3/28 Ab (middle panels in B and in part C ) or PMA and ionomycin (bottom panels in B ), and IL-2 production was assessed by intracellular staining as a function of GFP reporter gene expression ( B ) or by ELISA ( C ). In a separate approach, murine CD4 + CD25 neg T cells were transduced with Foxp3-IRES-GFP vector alone (x-axis of left panel in D) , NFAT2-IRES-NGFR alone (y-axis of left panel in D ), or Foxp3-IRES-GFP plus NFAT2-IRES-NGFR in combination (right panel in D ). In the left panel, singly transduced cells were mixed prior to flow cytometry to demonstrate that each population of cells could be spectrally separated. The transduced cells depicted in D were restimulated with plate-bound anti-CD3/28 Ab, and IL-2 production was measured by intracellular staining with an APC-conjugated Ab. The graph in E shows the mean fluorescence intensity (MFI) of IL-2 staining as a function of GFP (NFAT2) or NGFR (Empty vector, Foxp3, and Foxp3 + NFAT2) reporter MFI. Results are representative of 3 experiments.

    Article Snippet: Secretion of IL-2 into the culture supernatant by transduced cells (50,000 per well) activated for 8 hours on CD3/28-coated plates was measured using a standard IL-2 ELISA kit (eBioscience).

    Techniques: Expressing, Purification, Transduction, Plasmid Preparation, Staining, Cell Culture, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Cytometry, Fluorescence

    CD4+ CD25+ FoxP3+ classical regulatory T cells subset in not increased by DCs after efferocytosis. Top: Percentage of proliferating CFSE low , CD3 + , CD4 + ,CD25 + , FoxP3 + regulatory T cells in autologous T cell proliferation assays with immature DCs (iDCs, white circles), mature DCs (mDCs, white squares) and DCs loaded with apoptotic cells (NITApo-DCs, black circles) with insulin (20 µg/ml) at a ratio of 1∶10 for 7 days. Plots show the mean (line) of four independent experiments. Double-sided Wilcoxon test was used for the evaluation of statistical significance. Bottom: Representative FACS plots showing CD25 + FoxP3 + regulatory T cells gated on CFSE low , CD3 + , CD4 + .

    Journal: PLoS ONE

    Article Title: Efferocytosis Promotes Suppressive Effects on Dendritic Cells through Prostaglandin E2 Production in the Context of Autoimmunity

    doi: 10.1371/journal.pone.0063296

    Figure Lengend Snippet: CD4+ CD25+ FoxP3+ classical regulatory T cells subset in not increased by DCs after efferocytosis. Top: Percentage of proliferating CFSE low , CD3 + , CD4 + ,CD25 + , FoxP3 + regulatory T cells in autologous T cell proliferation assays with immature DCs (iDCs, white circles), mature DCs (mDCs, white squares) and DCs loaded with apoptotic cells (NITApo-DCs, black circles) with insulin (20 µg/ml) at a ratio of 1∶10 for 7 days. Plots show the mean (line) of four independent experiments. Double-sided Wilcoxon test was used for the evaluation of statistical significance. Bottom: Representative FACS plots showing CD25 + FoxP3 + regulatory T cells gated on CFSE low , CD3 + , CD4 + .

    Article Snippet: Quantitative PCR (qPCR) assays were performed under TaqMan universal assay conditions and using the following TaqMan Assays: Ccr7 (Mm01301785_m1), Ccl5 (Mm01302428_m1), Cd74 (Mm00658576_m1), Cd83 (Mm00486868_m1), Il2ra (Mm01340213_m1), Tnfrsf9 (Mm00441899_m1), Ins2 (Mm00731595_gH) and Iapp (Mm00439403_m1).

    Techniques: FACS